1 00:00:00,000 --> 00:00:02,415 [SQUEAKING] 2 00:00:02,415 --> 00:00:04,347 [RUSTLING] 3 00:00:04,347 --> 00:00:07,245 [CLICKING] 4 00:00:11,110 --> 00:00:12,940 PROFESSOR: Today I want to discuss 5 00:00:12,940 --> 00:00:14,590 nucleic acid metabolism. 6 00:00:14,590 --> 00:00:16,090 But before getting to that, I want 7 00:00:16,090 --> 00:00:19,300 to take a step back and talk a bit 8 00:00:19,300 --> 00:00:24,100 about what we closed with last time, which was discussing 9 00:00:24,100 --> 00:00:26,140 serine, glycine metabolism and how 10 00:00:26,140 --> 00:00:29,680 they can be used to generate one-carbon units that 11 00:00:29,680 --> 00:00:33,050 are for one-carbon transfer reactions. 12 00:00:33,050 --> 00:00:35,260 And these are one-carbon transfer reactions 13 00:00:35,260 --> 00:00:38,665 moving carbon that's more reduced than CO2. 14 00:00:38,665 --> 00:00:41,290 You'll remember, if we're going to do a carboxylation reaction, 15 00:00:41,290 --> 00:00:44,830 transfer of CO2, that often uses biotin, 16 00:00:44,830 --> 00:00:47,530 whereas if you're transferring a one-carbon unit that's 17 00:00:47,530 --> 00:00:51,310 more reduced than CO2, it comes from these folate species. 18 00:00:51,310 --> 00:00:54,040 And we described some of the different folate species-- 19 00:00:54,040 --> 00:00:58,420 N5-methyl-THF, which can donate a fully reduced 20 00:00:58,420 --> 00:01:01,060 one-carbon unit, a methyl group, to SAM, 21 00:01:01,060 --> 00:01:02,980 SAM being the universal methyl donor 22 00:01:02,980 --> 00:01:05,830 for most methylation reactions in cells. 23 00:01:05,830 --> 00:01:09,410 You can also have folates carry more oxidized one-carbon units, 24 00:01:09,410 --> 00:01:13,120 N5, N10-methylene-THF and N10-formyl-THF. 25 00:01:13,120 --> 00:01:15,130 And those will be very important today 26 00:01:15,130 --> 00:01:18,580 as we discuss one-carbon transfer reactions 27 00:01:18,580 --> 00:01:21,280 and nucleotide synthesis. 28 00:01:21,280 --> 00:01:23,710 Now, I recognize that folate metabolism 29 00:01:23,710 --> 00:01:26,050 can be very confusing. 30 00:01:26,050 --> 00:01:27,790 The structures are complex. 31 00:01:27,790 --> 00:01:31,480 The names are long and convoluted and hard 32 00:01:31,480 --> 00:01:34,420 to incorporate. 33 00:01:34,420 --> 00:01:37,750 But if you can think of it as just transferring 34 00:01:37,750 --> 00:01:40,870 one-carbon units of a different oxidation state, 35 00:01:40,870 --> 00:01:43,390 hopefully that will help you better understand what's 36 00:01:43,390 --> 00:01:46,460 going on in folate reactions. 37 00:01:46,460 --> 00:01:48,670 And so really, all it is, remember, serine 38 00:01:48,670 --> 00:01:50,230 has three carbons. 39 00:01:50,230 --> 00:01:52,460 Glycine has two carbons. 40 00:01:52,460 --> 00:01:55,360 And so when you convert serine to glycine, 41 00:01:55,360 --> 00:01:57,850 you end up generating a one-carbon unit. 42 00:01:57,850 --> 00:01:59,830 As I showed you in the lecture last time, 43 00:01:59,830 --> 00:02:06,040 this one-carbon unit is in the formaldehyde oxidation state. 44 00:02:06,040 --> 00:02:08,530 And so ultimately, that's generated. 45 00:02:08,530 --> 00:02:10,990 That's donated to a folate species 46 00:02:10,990 --> 00:02:22,760 to generate this carrier, the N5, N10-methylene-THF, 47 00:02:22,760 --> 00:02:30,770 which you can really think of as a carrier for one-carbon units 48 00:02:30,770 --> 00:02:33,890 in the formaldehyde oxidation state. 49 00:02:33,890 --> 00:02:39,440 Now, those can undergo an oxidation reaction. 50 00:02:39,440 --> 00:02:46,010 And so if we oxidize that carbon from the aldehyde to the acid, 51 00:02:46,010 --> 00:02:48,815 now we end up getting the N10-formyl-THF. 52 00:02:53,000 --> 00:02:58,640 All right, so that's carrying the one-carbon unit 53 00:02:58,640 --> 00:03:01,730 as the acid. 54 00:03:01,730 --> 00:03:09,570 Or we can reduce that one-carbon unit to the methyl group. 55 00:03:09,570 --> 00:03:17,490 And that's carrying it as the N5-methyl-THF, 56 00:03:17,490 --> 00:03:23,850 so carrying this as a methyl group, more reduced carbon 57 00:03:23,850 --> 00:03:25,170 than where we started. 58 00:03:25,170 --> 00:03:28,140 And so try to cut through all the names and whatnot 59 00:03:28,140 --> 00:03:32,220 and just realize that it's simply transfers of carbon 60 00:03:32,220 --> 00:03:33,960 in different oxidation states. 61 00:03:33,960 --> 00:03:35,760 And the relevant ones today is going 62 00:03:35,760 --> 00:03:39,810 to be N10-formyl and N5, N10-methylene-THF because it 63 00:03:39,810 --> 00:03:42,960 turns out generating those oxidized 64 00:03:42,960 --> 00:03:46,050 one-carbon units, formate groups, et cetera, 65 00:03:46,050 --> 00:03:50,040 is going to be important for purine as well 66 00:03:50,040 --> 00:03:52,170 as thymine synthesis. 67 00:03:52,170 --> 00:03:59,130 OK, so our topic today is really to discuss 68 00:03:59,130 --> 00:04:02,280 how we generate nucleic acids, how 69 00:04:02,280 --> 00:04:07,110 we break down purines and pyrimidines or purines. 70 00:04:07,110 --> 00:04:11,550 And this is really the final topic here in the course. 71 00:04:11,550 --> 00:04:16,950 In some ways, it's a bit fitting because it really incorporates 72 00:04:16,950 --> 00:04:21,930 lots of aspects of metabolism that we've been discussing 73 00:04:21,930 --> 00:04:24,930 over the course of the semester, all come together 74 00:04:24,930 --> 00:04:27,640 in nucleotide metabolism. 75 00:04:27,640 --> 00:04:32,850 Now, nucleotide metabolism itself is a very large topic, 76 00:04:32,850 --> 00:04:37,680 probably deserves more than one 90 minute lecture. 77 00:04:37,680 --> 00:04:40,110 But unfortunately, the time is what it is. 78 00:04:40,110 --> 00:04:43,410 And we just don't have time to cover every single detail. 79 00:04:43,410 --> 00:04:46,620 However, we can go pretty fast and at least 80 00:04:46,620 --> 00:04:49,110 give you a flavor of what's going on 81 00:04:49,110 --> 00:04:53,130 because as I've alluded to, this draws to lots of metabolism. 82 00:04:53,130 --> 00:04:56,130 I drew here just a very skeletonized view 83 00:04:56,130 --> 00:04:59,770 of glycolysis, the TCA cycle, the pentose phosphate pathway, 84 00:04:59,770 --> 00:05:03,810 couple aspects of amino acid synthesis, folate metabolism, 85 00:05:03,810 --> 00:05:06,330 and drew boxes around all the precursors 86 00:05:06,330 --> 00:05:09,150 that are necessary to generate various nucleotides. 87 00:05:09,150 --> 00:05:10,770 And you can see it's really drawn 88 00:05:10,770 --> 00:05:14,010 from across these metabolic pathways. 89 00:05:14,010 --> 00:05:16,080 As we go through these, you'll see 90 00:05:16,080 --> 00:05:18,240 that you actually know a lot. 91 00:05:18,240 --> 00:05:20,610 And this synthesizes, really, what 92 00:05:20,610 --> 00:05:23,610 we've learned from lots of other pathways coming together 93 00:05:23,610 --> 00:05:29,940 into understanding how cells make purines and pyrimidines. 94 00:05:29,940 --> 00:05:34,630 Now down here, I've color coded all the different-- 95 00:05:34,630 --> 00:05:35,610 so here's a pyrimidine. 96 00:05:35,610 --> 00:05:38,640 Here's a purine, the ring structure 97 00:05:38,640 --> 00:05:41,520 that you're probably familiar with from these purines 98 00:05:41,520 --> 00:05:43,950 and pyrimidines-- and color coded where 99 00:05:43,950 --> 00:05:46,770 it is that each of these atoms come from, where they're 100 00:05:46,770 --> 00:05:53,820 drawn from across metabolism. 101 00:05:53,820 --> 00:05:56,160 And again, it's a nice way to sort 102 00:05:56,160 --> 00:05:58,530 of synthesize lots of the pathways 103 00:05:58,530 --> 00:06:01,140 that we've discussed in class. 104 00:06:01,140 --> 00:06:05,700 Now ribose also is going to come from ribose 5-phosphate. 105 00:06:05,700 --> 00:06:08,730 That's, of course, a product of the pentose phosphate pathway, 106 00:06:08,730 --> 00:06:11,880 oxidative or non-oxidative, depending on how the cell wants 107 00:06:11,880 --> 00:06:13,420 to use those pathways-- 108 00:06:13,420 --> 00:06:15,420 the Calvin cycle and photosynthesis, 109 00:06:15,420 --> 00:06:18,270 lots of ways to get ribose 5-phosphate. 110 00:06:18,270 --> 00:06:20,220 That generates the ribose. 111 00:06:20,220 --> 00:06:22,890 And then there's a lot of complexity 112 00:06:22,890 --> 00:06:28,840 that goes into how one builds these complex base structures. 113 00:06:28,840 --> 00:06:33,720 Now, we just want to start here and show you on the slide here, 114 00:06:33,720 --> 00:06:37,350 here's a general purine base. 115 00:06:37,350 --> 00:06:40,080 Here's a general pyrimidine base. 116 00:06:40,080 --> 00:06:44,220 Kind of a stupid mnemonic that's always helped me 117 00:06:44,220 --> 00:06:47,430 that maybe will help you if you need to remember which 118 00:06:47,430 --> 00:06:50,160 of these is the purine and which of these is the pyrimidine 119 00:06:50,160 --> 00:06:55,150 as you go through, take MCAT exams or something like that-- 120 00:06:55,150 --> 00:06:58,890 so the purine ring looks kind of like a pope hat. 121 00:06:58,890 --> 00:07:00,480 And the pope is pure. 122 00:07:00,480 --> 00:07:02,310 And so it's kind of stupid, but it's 123 00:07:02,310 --> 00:07:05,730 one way to remember that this pope hat-looking structure is 124 00:07:05,730 --> 00:07:06,660 the purine. 125 00:07:06,660 --> 00:07:10,620 And the other one is the pyrimidine. 126 00:07:10,620 --> 00:07:13,560 Of course, you're familiar with purines and pyrimidines 127 00:07:13,560 --> 00:07:16,500 because these generate the various bases that 128 00:07:16,500 --> 00:07:19,470 are used for RNA and DNA, so adenine and guanine, the two 129 00:07:19,470 --> 00:07:22,980 purine bases in RNA and DNA, and then 130 00:07:22,980 --> 00:07:26,400 cytosine and uracil in RNA, and cytosine and thymine 131 00:07:26,400 --> 00:07:33,885 in DNA are the pyrimidine bases in RNA and DNA. 132 00:07:33,885 --> 00:07:37,478 Now, these are, of course, not the only possible purines 133 00:07:37,478 --> 00:07:38,145 and pyrimidines. 134 00:07:38,145 --> 00:07:40,080 They're not the only purines and pyrimidines 135 00:07:40,080 --> 00:07:42,230 that nature cares about. 136 00:07:42,230 --> 00:07:45,400 Here's a few other ones shown here. 137 00:07:45,400 --> 00:07:48,630 And so here is uric acid. 138 00:07:48,630 --> 00:07:52,680 Uric acid, as we've discussed, is the way birds 139 00:07:52,680 --> 00:07:55,200 excrete nitrogen. We'll discuss later today 140 00:07:55,200 --> 00:08:00,330 that it's the cause of gout, a very common human condition. 141 00:08:00,330 --> 00:08:04,250 There's also lots of other pharmacologically active 142 00:08:04,250 --> 00:08:06,450 molecules among the purines and pyrimidines. 143 00:08:06,450 --> 00:08:10,940 So here's caffeine, certainly a very important molecule 144 00:08:10,940 --> 00:08:15,110 for many of us to get going in the morning. 145 00:08:15,110 --> 00:08:17,750 There's things like adenine. 146 00:08:17,750 --> 00:08:21,830 So adenine plus the ribose sugar is a molecule called adenosine. 147 00:08:21,830 --> 00:08:24,410 Adenosine is a very important signaling molecule 148 00:08:24,410 --> 00:08:25,350 in our bodies. 149 00:08:25,350 --> 00:08:27,710 If you go to medical school, you'll 150 00:08:27,710 --> 00:08:32,690 use adenosine to figure out people's heart rhythms. 151 00:08:32,690 --> 00:08:34,471 There's also lots of drugs that come 152 00:08:34,471 --> 00:08:35,679 from purines and pyrimidines. 153 00:08:35,679 --> 00:08:36,990 So here's an example here. 154 00:08:36,990 --> 00:08:41,870 This is the drug 5-fluorouracil, a widely used cancer drug. 155 00:08:41,870 --> 00:08:44,870 And so it's an analog of a pyrimidine. 156 00:08:44,870 --> 00:08:49,610 Lots of anti-cancer drugs as well as antimicrobials 157 00:08:49,610 --> 00:08:53,270 end up being drugs that interfere 158 00:08:53,270 --> 00:08:57,800 with nucleotide metabolism and often are analogs 159 00:08:57,800 --> 00:09:00,920 of purines and pyrimidines. 160 00:09:00,920 --> 00:09:03,560 Now the first general comment I want to make 161 00:09:03,560 --> 00:09:06,620 is that in cells, the free bases of these things 162 00:09:06,620 --> 00:09:10,250 are much less useful than the nucleosides 163 00:09:10,250 --> 00:09:11,750 and the nucleotides. 164 00:09:11,750 --> 00:09:15,530 And so just to remind you of a little bit of nomenclature 165 00:09:15,530 --> 00:09:19,970 is that these purine and pyrimidine bases are usually 166 00:09:19,970 --> 00:09:25,480 attached to ribose and a nucleoside. 167 00:09:25,480 --> 00:09:30,560 So a nucleoside is basically the base, purine and pyrimidine 168 00:09:30,560 --> 00:09:33,440 base, plus ribose, but no phosphate. 169 00:09:33,440 --> 00:09:36,740 All right, so adenosine would be adenine stuck 170 00:09:36,740 --> 00:09:38,720 to ribose, but no phosphates on it. 171 00:09:38,720 --> 00:09:40,040 That's adenosine. 172 00:09:40,040 --> 00:09:43,730 And then you have your nucleotides. 173 00:09:43,730 --> 00:09:51,600 And the nucleotides are the base plus the ribose plus phosphate. 174 00:09:51,600 --> 00:09:55,100 And so it can be nucleotide monophosphate, diphosphate, 175 00:09:55,100 --> 00:09:58,790 triphosphate, AMP, ADP, ATP-- 176 00:09:58,790 --> 00:10:02,190 examples of nucleotides. 177 00:10:02,190 --> 00:10:09,710 Now, there are two major ways that cells build nucleotides. 178 00:10:09,710 --> 00:10:15,890 And that is they can make them de novo. 179 00:10:15,890 --> 00:10:20,420 So de novo is building them from scratch. 180 00:10:20,420 --> 00:10:27,890 Or you'll see that they can so-called salvage the bases. 181 00:10:27,890 --> 00:10:30,950 And this is, you might imagine, that if as animals, we 182 00:10:30,950 --> 00:10:32,000 eat other organisms. 183 00:10:32,000 --> 00:10:33,980 We take in their RNA and DNA. 184 00:10:33,980 --> 00:10:37,670 We get some of these purine and pyrimidine bases 185 00:10:37,670 --> 00:10:41,090 that these can be salvaged-- that is, added back 186 00:10:41,090 --> 00:10:46,340 to ribose to generate nucleosides and nucleotides. 187 00:10:46,340 --> 00:10:49,880 And you'll see, as we go through how you do de novo synthesis, 188 00:10:49,880 --> 00:10:51,980 that making these things is complex. 189 00:10:51,980 --> 00:10:54,540 It's energetically expensive. 190 00:10:54,540 --> 00:10:58,160 And so, of course, salvaging prevents 191 00:10:58,160 --> 00:11:02,000 you having to expend this energy and effort in order 192 00:11:02,000 --> 00:11:04,400 to generate them de novo. 193 00:11:04,400 --> 00:11:06,140 I also want to remind you because it's 194 00:11:06,140 --> 00:11:09,320 evident in the fact that uric acid is a way that birds 195 00:11:09,320 --> 00:11:12,920 and reptiles can excrete nitrogen is that just back 196 00:11:12,920 --> 00:11:17,480 to our prior discussion, that in the world, in the biosphere, 197 00:11:17,480 --> 00:11:21,050 nitrogen is a precious commodity. 198 00:11:21,050 --> 00:11:22,280 Remember, it has to be fixed. 199 00:11:22,280 --> 00:11:24,560 It's not infinitely available. 200 00:11:24,560 --> 00:11:29,210 Relatively few organisms can fix nitrogen into its useable form. 201 00:11:29,210 --> 00:11:34,820 And so salvaging premade bases is salvaging 202 00:11:34,820 --> 00:11:36,710 nitrogen in a usable form. 203 00:11:36,710 --> 00:11:39,300 And so evolutionarily, for a lot of organisms, 204 00:11:39,300 --> 00:11:42,500 that's also a beneficial thing to do. 205 00:11:42,500 --> 00:11:48,840 All right, now another last sort of introductory comment 206 00:11:48,840 --> 00:11:52,920 about why this lecture ends up being important and having 207 00:11:52,920 --> 00:11:55,260 some appreciation of these pathways 208 00:11:55,260 --> 00:11:57,720 is that certainly if you go to medical school, 209 00:11:57,720 --> 00:12:00,720 you will come across all kinds of things 210 00:12:00,720 --> 00:12:04,260 having to do with the metabolism of today's lecture. 211 00:12:04,260 --> 00:12:07,560 And that's because it turns out that making nucleosides 212 00:12:07,560 --> 00:12:10,110 and making nucleotides is limiting 213 00:12:10,110 --> 00:12:13,920 for proliferation in all kinds of biological contexts. 214 00:12:13,920 --> 00:12:16,140 And this is really back to what I alluded to earlier. 215 00:12:16,140 --> 00:12:18,480 A lot of our anti-cancer drugs, they also 216 00:12:18,480 --> 00:12:20,010 end up being anti-inflammatory. 217 00:12:20,010 --> 00:12:24,540 So things to treat inflammatory diseases and cancer oftentimes 218 00:12:24,540 --> 00:12:29,580 are things that block steps in nucleotide metabolism. 219 00:12:29,580 --> 00:12:32,910 Lots of antimicrobials or antibiotics 220 00:12:32,910 --> 00:12:36,330 also attack some of the same pathways, 221 00:12:36,330 --> 00:12:38,700 but are more specific for bacterial rather than 222 00:12:38,700 --> 00:12:40,740 human enzymes. 223 00:12:40,740 --> 00:12:45,750 And so really, these drugs converge on pathways 224 00:12:45,750 --> 00:12:48,840 that either produce or salvage nucleotides. 225 00:12:48,840 --> 00:12:52,770 And it's good to have some appreciation of where 226 00:12:52,770 --> 00:12:56,280 these things are acting because this ends up 227 00:12:56,280 --> 00:13:00,210 being really important for all kinds of aspects 228 00:13:00,210 --> 00:13:03,090 of human medicine, as well as understanding 229 00:13:03,090 --> 00:13:05,790 other bits of biology. 230 00:13:05,790 --> 00:13:11,160 All right, now either de novo synthesis 231 00:13:11,160 --> 00:13:14,820 of purine and pyrimidine nucleotides 232 00:13:14,820 --> 00:13:18,960 or salvage of those things, of purine and pyrimidine 233 00:13:18,960 --> 00:13:23,670 nucleotides, needs to add the base somehow to a ribose. 234 00:13:23,670 --> 00:13:29,100 And that starts with generating a molecule called PRPP, 235 00:13:29,100 --> 00:13:32,650 also abbreviated up there in my general overview. 236 00:13:32,650 --> 00:13:34,714 This stands for polyribo-- 237 00:13:41,260 --> 00:13:58,000 sorry, stands for phosphoribose pyrophosphate, PRPP, 238 00:13:58,000 --> 00:14:00,790 phosphoribose pyrophosphate. 239 00:14:00,790 --> 00:14:02,020 So what is that? 240 00:14:02,020 --> 00:14:12,340 Well, this is, if we draw here our ribose sugar and we draw it 241 00:14:12,340 --> 00:14:18,070 in the alpha-furanose form, so this would be 242 00:14:18,070 --> 00:14:24,700 alpha-5-ribose-phosphate so-- 243 00:14:27,520 --> 00:14:31,000 sorry, alpha-ribose 5-phosphate. 244 00:14:31,000 --> 00:14:34,380 OK, so that's ribose 5-phosphate. 245 00:14:34,380 --> 00:14:37,980 That's generated from pentose phosphate pathway, generated 246 00:14:37,980 --> 00:14:41,140 from the Calvin cycle, whatever. 247 00:14:41,140 --> 00:14:45,240 This is really starting point, discuss 248 00:14:45,240 --> 00:14:47,910 lots of ways we can generate that 249 00:14:47,910 --> 00:14:49,740 through different pathways. 250 00:14:49,740 --> 00:14:56,670 Well, if we take ATP and convert it to AMP-- 251 00:14:56,670 --> 00:14:58,570 that is, transfer two phosphates, 252 00:14:58,570 --> 00:15:01,830 pyrophosphorylate the one position-- 253 00:15:01,830 --> 00:15:04,620 now we end up with this molecule. 254 00:15:12,840 --> 00:15:19,650 OK, so this molecule here would be PRPP, 255 00:15:19,650 --> 00:15:22,860 so phosphoribose pyrophosphate. 256 00:15:22,860 --> 00:15:32,080 And PRPP can pick up a nitrogen, either the nitrogen 257 00:15:32,080 --> 00:15:35,830 from the base if it's the salvage pathway 258 00:15:35,830 --> 00:15:39,160 or whatever nitrogen-containing molecule 259 00:15:39,160 --> 00:15:41,770 is going to be the first step in de novo synthesis, which 260 00:15:41,770 --> 00:15:45,250 we'll get to later, and basically releases 261 00:15:45,250 --> 00:15:50,440 the pyrophosphate. 262 00:15:50,440 --> 00:15:54,430 That can be cleaved to two inorganic phosphates, 263 00:15:54,430 --> 00:15:56,230 pull the reaction forward. 264 00:15:56,230 --> 00:16:12,690 And what one ends up with is basically this nitrogen 265 00:16:12,690 --> 00:16:21,040 added either as the nucleoside monophosphate or some precursor 266 00:16:21,040 --> 00:16:25,030 to nitrogen-containing precursor to the nucleoside monophosphate 267 00:16:25,030 --> 00:16:27,970 that we can then build the purine or pyrimidine base 268 00:16:27,970 --> 00:16:30,410 on top. 269 00:16:30,410 --> 00:16:37,060 And so the dogma is that de novo synthesis of nucleotides 270 00:16:37,060 --> 00:16:40,360 is important for new proliferation of cells 271 00:16:40,360 --> 00:16:43,460 or a salvage is important for maintenance of cells. 272 00:16:43,460 --> 00:16:45,410 This is a huge oversimplification. 273 00:16:45,410 --> 00:16:49,390 It's effectively untrue as far as I'm concerned. 274 00:16:49,390 --> 00:16:51,910 Lots of drugs that affect salvage 275 00:16:51,910 --> 00:16:55,720 can affect diseases involved in proliferation and whatnot. 276 00:16:55,720 --> 00:17:00,370 But nonetheless, you should just realize that these early 277 00:17:00,370 --> 00:17:02,890 steps-- that is, adding PRPP-- 278 00:17:02,890 --> 00:17:05,170 is important either to salvage things 279 00:17:05,170 --> 00:17:07,960 or to do de novo synthesis. 280 00:17:07,960 --> 00:17:12,550 Now today, given our limited amount of time, 281 00:17:12,550 --> 00:17:16,113 I'm really going to focus on de novo synthesis of purines 282 00:17:16,113 --> 00:17:16,780 and pyrimidines. 283 00:17:16,780 --> 00:17:21,490 I will discuss a little bit about purine catabolism. 284 00:17:21,490 --> 00:17:25,000 Mainly this is because these are the most topics where there's 285 00:17:25,000 --> 00:17:27,609 something to say about biochemistry and metabolism 286 00:17:27,609 --> 00:17:29,800 that builds off a lot of the themes 287 00:17:29,800 --> 00:17:31,720 that we've covered in the class. 288 00:17:31,720 --> 00:17:35,737 They also focus on lots of aspects of relevant medicine. 289 00:17:35,737 --> 00:17:37,570 And so those of you who go to medical school 290 00:17:37,570 --> 00:17:38,470 will see this again. 291 00:17:38,470 --> 00:17:42,250 And I want you to appreciate it a little bit. 292 00:17:42,250 --> 00:17:44,920 And particularly, some aspects of salvage 293 00:17:44,920 --> 00:17:47,500 are important in biology and medicine. 294 00:17:47,500 --> 00:17:50,410 But really, my goal today is not for you 295 00:17:50,410 --> 00:17:56,750 to understand absolutely every chemical step that goes on, 296 00:17:56,750 --> 00:18:00,310 but really to appreciate how these things happen 297 00:18:00,310 --> 00:18:02,200 and hopefully synthesize some of what 298 00:18:02,200 --> 00:18:05,170 you've learned over the course of the semester. 299 00:18:05,170 --> 00:18:10,780 All right, so let's talk first about how you make purines, 300 00:18:10,780 --> 00:18:12,310 purine synthesis. 301 00:18:12,310 --> 00:18:16,750 So purine synthesis, you should know your MIT history. 302 00:18:16,750 --> 00:18:19,990 So the purine synthesis pathway, the de novo-- 303 00:18:19,990 --> 00:18:22,330 that is building purines from scratch-- 304 00:18:22,330 --> 00:18:26,110 was largely worked out by a guy by the name of Jack Buchanan. 305 00:18:26,110 --> 00:18:29,500 He partially did this at MIT. 306 00:18:29,500 --> 00:18:33,760 He basically was recruited from elsewhere, came to MIT. 307 00:18:33,760 --> 00:18:36,790 And when he was at MIT, also was the person 308 00:18:36,790 --> 00:18:39,790 who really modernized the MIT biology department 309 00:18:39,790 --> 00:18:42,200 to be what it is today. 310 00:18:42,200 --> 00:18:45,190 So prior to Jack Buchanan, biology department 311 00:18:45,190 --> 00:18:50,140 was much more about ecology and things like that. 312 00:18:50,140 --> 00:18:54,460 And really, he moved the MIT biology department 313 00:18:54,460 --> 00:18:57,770 into the molecular era. 314 00:18:57,770 --> 00:19:03,470 And so for the MIT students, it's good for you 315 00:19:03,470 --> 00:19:06,800 to appreciate some of our history. 316 00:19:06,800 --> 00:19:10,730 All right, now purine synthesis-- 317 00:19:10,730 --> 00:19:13,160 as I said, I'm going to go through all 318 00:19:13,160 --> 00:19:14,840 the steps of purine synthesis. 319 00:19:14,840 --> 00:19:17,990 You'll see it's complicated, all right? 320 00:19:17,990 --> 00:19:19,940 I will show you some of the chemistry. 321 00:19:19,940 --> 00:19:21,890 I don't have time to go into great detail 322 00:19:21,890 --> 00:19:24,170 or explain all of the steps in great detail. 323 00:19:24,170 --> 00:19:25,700 But again, my goal here is for you 324 00:19:25,700 --> 00:19:27,920 to appreciate how purines are made, 325 00:19:27,920 --> 00:19:33,380 not necessarily memorize all the steps in the pathway. 326 00:19:33,380 --> 00:19:37,250 All right, so purine synthesis starts 327 00:19:37,250 --> 00:19:43,590 with phosphoribose pyrophosphate, OK? 328 00:19:43,590 --> 00:19:48,030 We're going to build on the ribose base. 329 00:19:48,030 --> 00:19:54,750 And the first step is taking glutamine 330 00:19:54,750 --> 00:19:56,700 and converting it to glutamate. 331 00:19:56,700 --> 00:19:58,710 So what is that transferring? 332 00:19:58,710 --> 00:20:05,765 That's transferring a nitrogen. 333 00:20:05,765 --> 00:20:08,140 Remember, the difference between glutamine and glutamate, 334 00:20:08,140 --> 00:20:10,015 you can pick that up with glutamine synthase, 335 00:20:10,015 --> 00:20:15,340 is that was one way to put an ammonia onto glutamate to make 336 00:20:15,340 --> 00:20:16,300 glutamine. 337 00:20:16,300 --> 00:20:19,420 And so this ends up carrying out the reaction 338 00:20:19,420 --> 00:20:29,980 that I showed you earlier and adds 339 00:20:29,980 --> 00:20:37,270 a nitrogen to the ribose sugar, giving you this compound. 340 00:20:37,270 --> 00:20:43,930 All right, the next step is to take a glycine molecule. 341 00:20:43,930 --> 00:20:48,445 And that glycine molecule gets phosphorylated with ATP. 342 00:21:03,600 --> 00:21:08,370 So phosphorylate the carboxylic acid on the glycine molecule, 343 00:21:08,370 --> 00:21:26,710 and then this will combine with the prior molecule, 344 00:21:26,710 --> 00:21:31,900 releasing phosphate and generating this molecule, which 345 00:21:31,900 --> 00:21:37,720 I'm now going to just draw phosphoribose as like that 346 00:21:37,720 --> 00:21:41,425 because drawing ribose over and over again will become painful. 347 00:21:59,660 --> 00:22:02,640 And that generates this intermediate, 348 00:22:02,640 --> 00:22:12,204 which is called glycinamide ribonucleotide. 349 00:22:15,650 --> 00:22:17,660 I'm not going to write out nucleotide. 350 00:22:17,660 --> 00:22:22,130 And you will see that many of the steps in purine synthesis 351 00:22:22,130 --> 00:22:24,870 end up having these abbreviations. 352 00:22:24,870 --> 00:22:28,850 This one is called GAR for glycinamide ribonucleotide. 353 00:22:28,850 --> 00:22:31,400 And you'll see, as we get these long complex names, 354 00:22:31,400 --> 00:22:36,020 why one uses these abbreviations like GAR, which are actually 355 00:22:36,020 --> 00:22:39,740 used quite commonly. 356 00:22:39,740 --> 00:22:42,020 All right, the next step is we're 357 00:22:42,020 --> 00:22:47,970 going to add a one-carbon unit, a formate group. 358 00:22:47,970 --> 00:23:03,140 So remember, the formate carrier comes from N10-formyl-THF. 359 00:23:03,140 --> 00:23:10,960 And so-- eraser-- 360 00:23:22,850 --> 00:23:26,580 going to come from N10-formyl-THF, all right? 361 00:23:26,580 --> 00:23:37,690 So that's going to take the one-carbon unit off 362 00:23:37,690 --> 00:23:40,090 to regenerate THF, which, of course, can pick up 363 00:23:40,090 --> 00:23:43,240 the one-carbon unit again from serine and glycine, 364 00:23:43,240 --> 00:23:45,490 and then undergo an oxidation reaction to get 365 00:23:45,490 --> 00:23:48,970 the formate group, the N10-formyl-THF. 366 00:23:48,970 --> 00:24:23,380 And that generates this intermediate, 367 00:24:23,380 --> 00:24:32,910 which is called FGAR for formyl GAR, so formylglycinamide 368 00:24:32,910 --> 00:24:36,810 ribonucleotide, or FGAR. 369 00:24:36,810 --> 00:24:45,070 All right, the next step is we're going to add a nitro-- 370 00:24:45,070 --> 00:24:52,820 basically swap this oxygen here on this carbonyl to a nitrogen, 371 00:24:52,820 --> 00:24:56,470 so a C double bond to a nitrogen bond. 372 00:24:56,470 --> 00:24:59,230 And that requires-- so that's going 373 00:24:59,230 --> 00:25:02,260 to come from glutamine again. 374 00:25:02,260 --> 00:25:03,790 And it requires ATP. 375 00:25:06,980 --> 00:25:10,460 And it's somewhat of a complex reaction. 376 00:25:15,840 --> 00:25:17,340 And I think it's-- 377 00:25:20,580 --> 00:25:24,270 again, I will show you very briefly how this works. 378 00:25:24,270 --> 00:25:27,090 If you don't follow all of it, don't worry about it. 379 00:25:27,090 --> 00:25:33,030 But basically, if we just draw the middle part here 380 00:25:33,030 --> 00:25:37,750 of the molecule, I don't want to draw out the whole thing. 381 00:25:37,750 --> 00:25:42,220 So basically, this is phosphorylation here. 382 00:25:42,220 --> 00:25:45,720 So the phosphate from ATP transferred here 383 00:25:45,720 --> 00:25:48,120 to give this intermediate. 384 00:25:55,330 --> 00:26:17,300 That will lose a water, giving this intermediate. 385 00:26:17,300 --> 00:26:27,290 Then the ammonia from glutamine to glutamate 386 00:26:27,290 --> 00:26:31,940 come here, release the phosphate. 387 00:26:49,370 --> 00:26:53,570 And then typically, this is drawn where we just 388 00:26:53,570 --> 00:26:55,910 rearrange the double bond. 389 00:26:55,910 --> 00:27:11,860 And one ends up with this intermediate, 390 00:27:11,860 --> 00:27:13,020 which is called FGAM. 391 00:27:57,860 --> 00:28:00,050 So this is this intermediate, which 392 00:28:00,050 --> 00:28:20,000 is FGAM for formyl glycinamide ribonucleotide, so FGAM. 393 00:28:20,000 --> 00:28:24,130 All right, makes sense? 394 00:28:24,130 --> 00:28:32,110 All right, the next reaction is also a little bit complex 395 00:28:32,110 --> 00:28:34,450 and maybe hard to see. 396 00:28:34,450 --> 00:28:35,890 Again, I will draw it out. 397 00:28:35,890 --> 00:28:38,320 If you don't follow all of it, don't worry about it. 398 00:28:38,320 --> 00:28:40,780 But it's nice for those of you who 399 00:28:40,780 --> 00:28:43,340 want to follow it to see what's happening. 400 00:28:43,340 --> 00:28:46,990 So again, now we're going to take another ATP, 401 00:28:46,990 --> 00:28:53,380 hydrolyze it to ADP plus Pi, release of water. 402 00:28:53,380 --> 00:28:55,630 And in the end, what we're going to generate 403 00:28:55,630 --> 00:29:40,570 is we're going to close this ring to generate-- so we're 404 00:29:40,570 --> 00:29:43,390 going to close that ring, so make 405 00:29:43,390 --> 00:29:47,050 a bond from this nitrogen to this carbon, close this ring. 406 00:29:47,050 --> 00:29:56,076 This generates this intermediate called AIR, or amino ribo-- 407 00:29:56,076 --> 00:30:03,288 sorry, aminoimidazole ribonucleotide. 408 00:30:06,030 --> 00:30:07,530 Don't worry about the long names. 409 00:30:07,530 --> 00:30:09,930 Everyone would just call it AIR. 410 00:30:09,930 --> 00:30:11,980 So how does this happen? 411 00:30:11,980 --> 00:30:13,530 Well, it happens as follows. 412 00:30:29,130 --> 00:30:37,820 So first step is phosphorylation of-- 413 00:30:37,820 --> 00:30:41,120 so ATP will phosphorylate the aldehyde 414 00:30:41,120 --> 00:30:43,980 to give this intermediate. 415 00:30:48,580 --> 00:30:49,420 Water's lost. 416 00:31:01,670 --> 00:31:10,755 OK, now you have this nitrogen down here, can-- 417 00:31:16,260 --> 00:31:18,030 losing the phosphate. 418 00:31:20,630 --> 00:31:31,710 OK, so that will end up closing the ring 419 00:31:31,710 --> 00:31:34,320 on that side of the molecule. 420 00:31:34,320 --> 00:31:39,870 And then all I've done is I've just removed the electrons 421 00:31:39,870 --> 00:31:43,620 here so that rather than being a carbon-nitrogen double bond, 422 00:31:43,620 --> 00:31:46,680 it's now a carbon-carbon double bond. 423 00:31:46,680 --> 00:31:51,990 And that generates FGAM to AIR, costing an ATP-- 424 00:31:51,990 --> 00:31:56,900 again, one of the more complicated reactions to see. 425 00:31:56,900 --> 00:31:58,805 Again, if you're into how the reactions work, 426 00:31:58,805 --> 00:32:02,270 hopefully I gave you enough information to understand it. 427 00:32:02,270 --> 00:32:07,810 If you're not, just realize that this is the next step. 428 00:32:07,810 --> 00:32:14,320 OK, the next step is a reaction that's very similar to one 429 00:32:14,320 --> 00:32:16,960 that we saw in the urea cycle. 430 00:32:16,960 --> 00:32:21,160 And that is we are going to add-- 431 00:32:21,160 --> 00:32:22,210 oh, sorry. 432 00:32:22,210 --> 00:32:27,010 The next step is one that-- 433 00:32:27,010 --> 00:32:27,560 sorry. 434 00:32:27,560 --> 00:32:29,140 Before we do that step, the next step 435 00:32:29,140 --> 00:32:34,370 is one where we're going to add a CO2 group to this carbon. 436 00:32:34,370 --> 00:32:37,000 So that's a carboxylation reaction. 437 00:32:37,000 --> 00:32:40,300 You would typically think of a carboxylation reaction 438 00:32:40,300 --> 00:32:42,490 to involve biotin. 439 00:32:42,490 --> 00:32:44,860 This is one that doesn't. 440 00:32:44,860 --> 00:32:48,730 It's an exception to that rule, does not involve biotin. 441 00:32:48,730 --> 00:32:50,825 But it does involve ATP. 442 00:32:55,090 --> 00:33:02,200 OK, so remember, the way we did this before is we 443 00:33:02,200 --> 00:33:07,420 would have phosphorylation of our bicarbonate. 444 00:33:07,420 --> 00:33:11,050 So use ATP to phosphorylate bicarbonate. 445 00:33:11,050 --> 00:33:32,460 And so if we just draw this part of the molecule here, 446 00:33:32,460 --> 00:33:36,510 displacing the phosphate, and then what we end up with is-- 447 00:33:51,030 --> 00:33:53,020 OK, we end up with that. 448 00:33:53,020 --> 00:33:55,650 And if I just rearrange so that the double bond is here 449 00:33:55,650 --> 00:34:34,270 instead of there, then we end up with this next intermediate, 450 00:34:34,270 --> 00:34:49,760 which is CAIR or carboxy-AIR, so CAIR or carboxy-AIR. 451 00:34:49,760 --> 00:34:51,960 All right, now we get to the step 452 00:34:51,960 --> 00:34:56,760 that's very similar to the one that we saw in the urea cycle 453 00:34:56,760 --> 00:34:59,880 where we're going to use aspartate to fumerate 454 00:34:59,880 --> 00:35:06,550 conversion to add a nitrogen to this carboxylic acid. 455 00:35:06,550 --> 00:35:16,930 And so you'll remember that way we did that in the urea cycle 456 00:35:16,930 --> 00:35:23,380 is we used ATP to AMP conversion to end up 457 00:35:23,380 --> 00:35:25,900 adding that aspartate group. 458 00:35:25,900 --> 00:35:29,650 In this case, it's just ATP to ADP plus Pi. 459 00:35:29,650 --> 00:35:41,760 OK, so that's going to phosphorylate this acid group. 460 00:35:41,760 --> 00:35:43,830 OK, so I'm not going to draw the whole thing. 461 00:35:43,830 --> 00:35:45,500 Phosphorylate that, and then we'll 462 00:35:45,500 --> 00:35:49,880 have an aspartate molecule. 463 00:36:07,360 --> 00:36:12,070 OK, so this is the amino acid aspartate. 464 00:36:17,962 --> 00:36:20,350 OK, and one gets-- 465 00:37:17,930 --> 00:37:23,330 didn't leave myself enough room, but that's 466 00:37:23,330 --> 00:37:32,740 an aspartate molecule bound up there to that carboxylic acid, 467 00:37:32,740 --> 00:37:33,950 just like we saw before. 468 00:37:33,950 --> 00:37:38,200 This is a molecule called SAICAR, 469 00:37:38,200 --> 00:37:40,720 which is succinyl amino imidazole 470 00:37:40,720 --> 00:38:01,090 carboxy amide ribonucleotide. 471 00:38:01,090 --> 00:38:05,410 OK, definitely everyone calls it SAICAR. 472 00:38:05,410 --> 00:38:08,650 Now just as we saw then from the urea cycle, 473 00:38:08,650 --> 00:38:13,620 we are now going to take off a fumerate. 474 00:38:19,060 --> 00:38:24,220 OK, so this is reaction that you've seen before. 475 00:38:33,400 --> 00:38:37,840 OK, just like we saw in the urea cycle, so that reaction, 476 00:38:37,840 --> 00:38:39,280 lose the fumerate. 477 00:38:39,280 --> 00:38:42,190 And what you're left with is basically 478 00:38:42,190 --> 00:38:45,715 just the nitrogen left behind. 479 00:39:15,440 --> 00:39:22,830 OK, and you get this intermediate, 480 00:39:22,830 --> 00:39:27,860 which is referred to as AICAR, so 481 00:39:27,860 --> 00:39:31,820 amino-imidazole-carboxy-amide ribonucleotide, 482 00:39:31,820 --> 00:39:36,670 so minus the succinate group, AICAR. 483 00:39:36,670 --> 00:39:42,700 OK, next step is we're going to use a formate group 484 00:39:42,700 --> 00:39:51,690 and add it to this nitrogen. So that's straightforward. 485 00:39:51,690 --> 00:39:55,440 So if we draw out here our formate group 486 00:39:55,440 --> 00:40:17,750 from N10-formyl-THF, so that will be our N10-formyl-THF, 487 00:40:17,750 --> 00:40:19,880 generate THF. 488 00:40:19,880 --> 00:40:21,425 And then we get-- 489 00:41:01,360 --> 00:41:08,870 OK, let me get that formate group added there. 490 00:41:08,870 --> 00:41:18,400 And so this is FAICAR, or formyl AICAR, FAICAR. 491 00:41:18,400 --> 00:41:24,210 All right, next step is we're going to close the ring. 492 00:41:30,730 --> 00:41:35,995 And so close the ring, that's going to give this. 493 00:41:45,710 --> 00:41:50,350 OK, so that part of the molecule where the ring closes, 494 00:41:50,350 --> 00:41:55,480 it just rearrange like that to lose water. 495 00:42:05,550 --> 00:43:15,580 And that is going to generate our purine ring the way 496 00:43:15,580 --> 00:43:19,150 you're used to seeing it drawn. 497 00:43:19,150 --> 00:43:29,170 And so this is the purine IMP for inosine monophosphate. 498 00:43:29,170 --> 00:43:36,730 And inosine monophosphate is the precursor for all purines. 499 00:43:36,730 --> 00:43:40,220 And so if you look at this, what you see is that in the end, 500 00:43:40,220 --> 00:43:45,040 there is two nitrogens that come from ammonia, from glutamine 501 00:43:45,040 --> 00:43:47,470 to glutamate, from the ammonia released from glutamine 502 00:43:47,470 --> 00:43:51,130 to glutamate, one nitrogen here that comes from glycine, 503 00:43:51,130 --> 00:43:54,160 and that nitrogen there that comes from aspartate. 504 00:43:54,160 --> 00:43:58,420 Two of the carbons, these two, come from the formate group, 505 00:43:58,420 --> 00:44:01,030 from N10-formyl-THF. 506 00:44:01,030 --> 00:44:04,840 Another carbon up here comes from CO2. 507 00:44:04,840 --> 00:44:06,880 And the remaining two carbons here 508 00:44:06,880 --> 00:44:10,370 come from a glycine molecule, with, of course, 509 00:44:10,370 --> 00:44:14,020 the ribose phosphate from PRPP. 510 00:44:14,020 --> 00:44:16,150 And so if you go through, what you'll find 511 00:44:16,150 --> 00:44:18,820 is that it took two ATP to generate 512 00:44:18,820 --> 00:44:22,330 the PRPP from ribose phosphate-- so starting 513 00:44:22,330 --> 00:44:27,400 with a ribose phosphate, two ATP to generate the PRPP, and then 514 00:44:27,400 --> 00:44:28,922 five additional ATPs. 515 00:44:28,922 --> 00:44:30,880 So if you go up and look through all the steps, 516 00:44:30,880 --> 00:44:34,150 there are five steps that require ATP, so quite 517 00:44:34,150 --> 00:44:38,410 a lot of ATP to generate this purine ring, 518 00:44:38,410 --> 00:44:41,080 inosine monophosphate. 519 00:44:41,080 --> 00:44:45,610 Now, inosine is not found in RNA or DNA. 520 00:44:45,610 --> 00:44:47,980 And so this has to be, of course, turned 521 00:44:47,980 --> 00:44:49,690 into adenine and guanine. 522 00:44:49,690 --> 00:44:51,890 And so how does that happen? 523 00:44:51,890 --> 00:44:52,900 Well, I'll show you. 524 00:44:52,900 --> 00:44:59,900 And so turning it, inosine monophosphate into adenine, 525 00:44:59,900 --> 00:45:04,790 basically involves putting a nitrogen here 526 00:45:04,790 --> 00:45:07,220 instead of this double bond to oxygen. 527 00:45:07,220 --> 00:45:11,570 And so that occurs via a very similar-- 528 00:45:11,570 --> 00:45:14,000 in fact, the same reaction, that we 529 00:45:14,000 --> 00:45:21,560 saw to take CAIR and generate SAICAR, all right? 530 00:45:21,560 --> 00:45:25,910 And so it was basically using aspartate to transfer 531 00:45:25,910 --> 00:45:28,283 that nitrogen. I don't have time to go 532 00:45:28,283 --> 00:45:29,450 through the whole mechanism. 533 00:45:29,450 --> 00:45:34,230 But it's basically what I already showed you. 534 00:45:34,230 --> 00:45:42,740 And so you take GTP plus acetate, get GDP plus Pi. 535 00:45:42,740 --> 00:45:46,430 Note, you use guanine triphosphate, 536 00:45:46,430 --> 00:45:49,370 so GTP rather than ATP, to end up generating 537 00:45:49,370 --> 00:45:51,530 the adenine nucleotide. 538 00:45:51,530 --> 00:45:55,370 All right, it's going to release water. 539 00:45:55,370 --> 00:46:00,050 Water was also released in that reaction. 540 00:46:00,050 --> 00:46:03,710 And what you end up with is this intermediate. 541 00:46:03,710 --> 00:46:05,750 So I'm not going to draw the whole structure 542 00:46:05,750 --> 00:46:08,090 for the sake of time. 543 00:46:08,090 --> 00:46:14,710 But this here would be the top here of the purine ring. 544 00:46:14,710 --> 00:46:16,215 So this would have a-- 545 00:46:26,820 --> 00:46:32,910 so that's going to add an aspartate molecule up there. 546 00:46:32,910 --> 00:46:39,060 And then from that, we're going to release fumerate. 547 00:46:47,700 --> 00:46:49,770 OK, so fumerate comes off. 548 00:46:52,490 --> 00:46:54,725 And what we're left with is-- 549 00:47:19,780 --> 00:47:26,320 OK, and this here is adenosine monophosphate, AMP. 550 00:47:26,320 --> 00:47:31,440 And, of course, I can just take two phosphorylation events 551 00:47:31,440 --> 00:47:32,635 and get ATP. 552 00:47:36,030 --> 00:47:39,900 Basically, phospho transfer ATP to make ADP, 553 00:47:39,900 --> 00:47:42,390 and then, ultimately, ADP to make 554 00:47:42,390 --> 00:47:46,540 ATP via glycolysis, oxidative phosphorylation, whatever. 555 00:47:46,540 --> 00:47:51,470 And so that's how you generate adenine nucleotides. 556 00:47:51,470 --> 00:47:56,060 Generate guanine nucleotides, what has to happen 557 00:47:56,060 --> 00:48:01,460 is that we need to put a nitrogen here. 558 00:48:01,460 --> 00:48:05,990 OK, so putting a nitrogen on this carbon, 559 00:48:05,990 --> 00:48:10,520 first thing that happens is we add water 560 00:48:10,520 --> 00:48:12,530 across that double bond. 561 00:48:24,510 --> 00:48:27,750 OK, so that's this part here of the molecule, 562 00:48:27,750 --> 00:48:32,010 added water across that double bond. 563 00:48:32,010 --> 00:48:37,770 Now we're going to oxidize this carbon-nitrogen bond, 564 00:48:37,770 --> 00:48:40,155 seen this a million times-- 565 00:48:42,890 --> 00:48:44,900 generates a hydride ion. 566 00:48:44,900 --> 00:48:47,540 What's that hydride ion-- or sorry, 567 00:48:47,540 --> 00:48:52,830 oxidize this alcohol to the ketone, 568 00:48:52,830 --> 00:48:54,860 so generates a hydride ion. 569 00:48:54,860 --> 00:49:00,740 Those electrons go to NAD+, generate an NADH, 570 00:49:00,740 --> 00:49:02,630 oxidation reaction. 571 00:49:02,630 --> 00:49:04,505 And that gives us this. 572 00:49:17,730 --> 00:49:22,390 OK, and then the next step is that we're 573 00:49:22,390 --> 00:49:26,230 going to take that ketone that was generated here, 574 00:49:26,230 --> 00:49:30,070 and we're going to turn it into, basically, 575 00:49:30,070 --> 00:49:33,010 replace that oxygen with a nitrogen. 576 00:49:33,010 --> 00:49:36,100 And so that's exactly the reaction that we saw when 577 00:49:36,100 --> 00:49:40,000 we went here from FGAR to FGAM. 578 00:49:40,000 --> 00:49:43,420 All right, so if I go from same reaction 579 00:49:43,420 --> 00:49:48,230 that I used to go from FGAR to FGAM, 580 00:49:48,230 --> 00:49:56,410 so this is ATP goes to ADP plus Pi. 581 00:49:56,410 --> 00:50:02,440 All right, nitrogen comes here from glutamine 582 00:50:02,440 --> 00:50:08,780 to glutamate, same reaction that occurs there. 583 00:50:08,780 --> 00:50:10,450 Water comes off. 584 00:50:10,450 --> 00:50:15,760 What I end up doing is I replace this. 585 00:50:15,760 --> 00:50:19,800 I'll end up getting this structure instead. 586 00:50:19,800 --> 00:50:21,900 And then I can just rearrange the double bond 587 00:50:21,900 --> 00:50:26,310 to be on the ring rather than to the nitrogen outside the ring. 588 00:50:26,310 --> 00:50:27,510 And that generates-- 589 00:50:51,660 --> 00:50:56,690 OK, and so this is GMP. 590 00:50:56,690 --> 00:51:01,970 Notice that the ATP was used to make GMP. 591 00:51:01,970 --> 00:51:04,790 GTP was used to make AMP. 592 00:51:04,790 --> 00:51:09,110 So use the opposite purine to make each reaction. 593 00:51:09,110 --> 00:51:11,150 I know I went through these quickly, 594 00:51:11,150 --> 00:51:13,550 just don't have time to go into it in more detail. 595 00:51:13,550 --> 00:51:17,300 But it's really just repurposing similar reactions 596 00:51:17,300 --> 00:51:19,250 that are involved in purine synthesis 597 00:51:19,250 --> 00:51:22,520 to end up changing this inosine monophosphate 598 00:51:22,520 --> 00:51:26,390 into AMP or into GMP. 599 00:51:26,390 --> 00:51:28,730 And I guess for consistency, I should 600 00:51:28,730 --> 00:51:31,430 have drawn that in yellow because the nitrogen comes 601 00:51:31,430 --> 00:51:34,070 from the glutamine. 602 00:51:34,070 --> 00:51:36,980 OK, so that is purine synthesis. 603 00:51:36,980 --> 00:51:40,010 That's how you get AMP and GMP. 604 00:51:40,010 --> 00:51:43,700 If you're going to break these things down, 605 00:51:43,700 --> 00:51:47,990 of course, you're going to use separate pathways. 606 00:51:47,990 --> 00:51:49,940 Why are you going to use separate pathways? 607 00:51:49,940 --> 00:51:53,150 Well, because as we've heard over and over again 608 00:51:53,150 --> 00:51:55,670 in the class, for thermodynamic reasons, 609 00:51:55,670 --> 00:51:58,790 delta G has to be less than 0 for any pathway to work, 610 00:51:58,790 --> 00:52:00,710 can't be less than 0 in both directions. 611 00:52:00,710 --> 00:52:02,510 One direction needs energy input. 612 00:52:02,510 --> 00:52:07,550 I had energy input to make these purine nucleotides. 613 00:52:07,550 --> 00:52:11,330 Opposite direction don't need energy to make it. 614 00:52:11,330 --> 00:52:14,210 I don't have time to get into all the details 615 00:52:14,210 --> 00:52:15,680 of purine breakdown. 616 00:52:15,680 --> 00:52:18,140 Although, here are some things. 617 00:52:18,140 --> 00:52:22,560 I just summarize it here for you on this slide. 618 00:52:22,560 --> 00:52:26,090 Basically, you start from AMP. 619 00:52:26,090 --> 00:52:27,410 You take the phosphate off. 620 00:52:27,410 --> 00:52:28,970 You end up generating adenosine. 621 00:52:28,970 --> 00:52:33,110 You end up going to inosine, remove the ribose. 622 00:52:33,110 --> 00:52:35,360 Then you get another base. 623 00:52:35,360 --> 00:52:38,390 And the base on inosine monophosphate, 624 00:52:38,390 --> 00:52:42,980 when it's just the base, so the nucleoside and nucleotide is 625 00:52:42,980 --> 00:52:45,890 inosine, when you get rid of the ribose, 626 00:52:45,890 --> 00:52:49,790 now the base is called hypoxanthine. 627 00:52:49,790 --> 00:52:52,880 It's a little bit of a confusing thing. 628 00:52:52,880 --> 00:52:55,100 But just recognize that hypoxanthine 629 00:52:55,100 --> 00:52:58,680 is the base that is found on inosine or inosine 630 00:52:58,680 --> 00:53:00,500 monophosphate, all right? 631 00:53:00,500 --> 00:53:04,770 And so just to summarize what I showed 632 00:53:04,770 --> 00:53:10,080 you what was on the slide, so to break down AMP, 633 00:53:10,080 --> 00:53:12,270 you'd remove the phosphate. 634 00:53:12,270 --> 00:53:15,960 Now you end up with the nucleoside adenosine. 635 00:53:15,960 --> 00:53:20,795 So that's adenine plus the ribose. 636 00:53:24,560 --> 00:53:30,860 Take off the nitrogen to move back to the nucleoside inosine. 637 00:53:30,860 --> 00:53:34,970 Now we can remove the ribose from inosine. 638 00:53:34,970 --> 00:53:38,210 What we're left with is the base, which is hypoxanthine. 639 00:53:40,740 --> 00:53:45,050 OK, so I'll draw out hypoxanthine for you here. 640 00:53:56,740 --> 00:54:01,030 OK, so hypoxanthine, you'll see is basically 641 00:54:01,030 --> 00:54:04,690 exactly what we drew out for inosine monophosphate. 642 00:54:04,690 --> 00:54:09,790 It's just the base from inosine monophosphate is hypoxanthine. 643 00:54:09,790 --> 00:54:15,550 Now if we oxidize this carbon-nitrogen bond, which 644 00:54:15,550 --> 00:54:20,740 is carried out by a complex oxidation reaction, 645 00:54:20,740 --> 00:54:27,730 requiring oxygen and water and producing hydrogen peroxide 646 00:54:27,730 --> 00:54:35,480 by an enzyme called xanthine oxidase. 647 00:54:43,810 --> 00:54:47,800 OK, so the enzyme xanthine oxidase-- interesting enzyme-- 648 00:54:47,800 --> 00:54:50,110 has a molybdenum group. 649 00:54:50,110 --> 00:54:53,215 And in it, is a cofactor and iron and FAD-- 650 00:54:53,215 --> 00:54:56,680 a complex redox reaction we don't have time to get into-- 651 00:54:56,680 --> 00:55:01,600 ends up generating this purine base. 652 00:55:17,340 --> 00:55:25,420 OK, this purine base is xanthine. 653 00:55:25,420 --> 00:55:36,940 Xanthine oxidase, same enzyme, can also 654 00:55:36,940 --> 00:55:40,690 use the same chemistry to oxidize 655 00:55:40,690 --> 00:55:43,080 this nitrogen-carbon bond. 656 00:55:43,080 --> 00:55:47,140 So this one here, when it was oxidized there, 657 00:55:47,140 --> 00:55:49,700 now we're going to oxidize this one here. 658 00:55:49,700 --> 00:55:51,340 And what does that generate? 659 00:56:00,830 --> 00:56:12,610 That generates this compound, uric acid. 660 00:56:12,610 --> 00:56:16,080 And so for the birds and reptiles to excrete nitrogen, 661 00:56:16,080 --> 00:56:20,190 it would synthesize a purine like AMP, 662 00:56:20,190 --> 00:56:22,680 and then break it down to uric acid 663 00:56:22,680 --> 00:56:28,320 as a way to eliminate nitrogen, all right? 664 00:56:28,320 --> 00:56:36,220 Guanine GMP, lose a phosphate, you 665 00:56:36,220 --> 00:56:42,550 get the nucleoside guanosine, so guanine the base, 666 00:56:42,550 --> 00:56:45,610 plus the ribose with no phosphate. 667 00:56:45,610 --> 00:56:49,070 Take the ribose group off. 668 00:56:49,070 --> 00:56:50,315 Now we're left with guanine. 669 00:56:53,450 --> 00:57:01,430 And then if we start with guanine, 670 00:57:01,430 --> 00:57:08,210 remove nitrogen from guanine, then we end up with xanthine. 671 00:57:08,210 --> 00:57:11,840 OK, so remember, there was additional oxidation step 672 00:57:11,840 --> 00:57:14,780 to make GMP that wasn't there in AMP. 673 00:57:14,780 --> 00:57:16,850 That means we get a more oxidized product back 674 00:57:16,850 --> 00:57:17,790 when we break it down. 675 00:57:17,790 --> 00:57:19,400 So it's broken down to xanthine. 676 00:57:19,400 --> 00:57:23,820 And that can be acted on also to generate uric acid. 677 00:57:23,820 --> 00:57:26,090 And so you could also make GMP, and then 678 00:57:26,090 --> 00:57:28,460 break down the GMP to make uric acid as a way 679 00:57:28,460 --> 00:57:31,740 to excrete nitrogen. 680 00:57:31,740 --> 00:57:34,840 Now uric acid is also medically important in humans. 681 00:57:34,840 --> 00:57:37,840 This is, of course, the thing that is the cause of gout. 682 00:57:37,840 --> 00:57:42,540 So gout is uric acid, is produced in excess, deposits 683 00:57:42,540 --> 00:57:45,850 and crystallizes in joints and causes a lot of pain. 684 00:57:45,850 --> 00:57:49,380 And one way to treat gout is with a xanthine oxidase 685 00:57:49,380 --> 00:57:49,990 inhibitor. 686 00:57:49,990 --> 00:57:51,720 There's a drug called allopurinol 687 00:57:51,720 --> 00:57:54,300 that inhibits xanthine oxidase. 688 00:57:54,300 --> 00:57:58,410 And that ends up being a way to slow 689 00:57:58,410 --> 00:58:02,920 the production of uric acid and can treat patients with gout. 690 00:58:02,920 --> 00:58:08,860 All right, now, if you break down purines 691 00:58:08,860 --> 00:58:14,980 and you want to recapture them, you can also use salvage. 692 00:58:14,980 --> 00:58:19,140 And so to save some time, here I drew out, 693 00:58:19,140 --> 00:58:22,380 basically, the salvage pathways for purines. 694 00:58:22,380 --> 00:58:24,120 And so you can start with adenine, 695 00:58:24,120 --> 00:58:25,800 or you can start with guanine. 696 00:58:25,800 --> 00:58:27,990 Or you can start with hypoxanthine. 697 00:58:27,990 --> 00:58:31,980 And basically, these are simply added to PRPP. 698 00:58:31,980 --> 00:58:35,870 So if you take adenine, add it to PRPP, now you have AMP. 699 00:58:35,870 --> 00:58:40,500 If you take hypoxanthine, add it to PRPP, now you have IMP. 700 00:58:40,500 --> 00:58:43,500 And you can use that IMP to then synthesize AMP, 701 00:58:43,500 --> 00:58:48,240 or not shown here, GMP, by the pathways I've already shown. 702 00:58:48,240 --> 00:58:50,850 Or if you have guanine, you can salvage that guanine, 703 00:58:50,850 --> 00:58:55,230 add it to PRPP, and then generate GMP. 704 00:58:55,230 --> 00:58:57,405 And so different enzymes do this. 705 00:58:57,405 --> 00:59:00,210 This enzyme here, hypoxanthine-guanine 706 00:59:00,210 --> 00:59:01,980 phosphoribosyltransferase, which is 707 00:59:01,980 --> 00:59:04,260 how you take guanine or hypoxanthine 708 00:59:04,260 --> 00:59:07,860 and make IMP or GMP via salvage pathway 709 00:59:07,860 --> 00:59:11,970 is actually a very famous enzyme because it's 710 00:59:11,970 --> 00:59:14,310 part of an inborn area of metabolism called 711 00:59:14,310 --> 00:59:17,790 Lesch-Nyhan syndrome or hypoguanine 712 00:59:17,790 --> 00:59:22,130 phosphoribosyltransferase or HPRT. 713 00:59:25,850 --> 00:59:32,685 So HPRT deficiency is what causes Lesch-Nyhan syndrome. 714 00:59:36,200 --> 00:59:38,380 And it's an inborn error of metabolism. 715 00:59:38,380 --> 00:59:42,700 You're deficient in this HGPRT enzyme, 716 00:59:42,700 --> 00:59:46,150 which means you can't salvage guanine or hypoxanthine. 717 00:59:46,150 --> 00:59:50,080 Now I bring this up, not because we have time to talk about all 718 00:59:50,080 --> 00:59:53,170 of the inborn errors of metabolism, 719 00:59:53,170 --> 00:59:57,070 but mentioning this one really points out 720 00:59:57,070 --> 00:59:58,660 that we don't understand metabolism 721 00:59:58,660 --> 01:00:00,410 as well as we think we do. 722 01:00:00,410 --> 01:00:01,990 So if I ask you to predict what would 723 01:00:01,990 --> 01:00:03,940 be the consequences of an inability 724 01:00:03,940 --> 01:00:06,460 to salvage guanine and hypoxanthine, 725 01:00:06,460 --> 01:00:09,550 you might say, oh, well, that means 726 01:00:09,550 --> 01:00:12,160 that you're going to not be able to take these things up. 727 01:00:12,160 --> 01:00:16,330 Maybe you'll have excess guanine and hypoxanthine breakdown. 728 01:00:16,330 --> 01:00:19,960 And maybe you'll get more uric acid and get gout. 729 01:00:19,960 --> 01:00:21,370 And that's absolutely the case. 730 01:00:21,370 --> 01:00:25,510 Patients with Lesch-Nyhan syndrome, HPRT deficiency, 731 01:00:25,510 --> 01:00:27,010 do indeed get gout. 732 01:00:27,010 --> 01:00:28,430 That makes sense. 733 01:00:28,430 --> 01:00:30,040 But the main problem they have is they 734 01:00:30,040 --> 01:00:32,260 have a self mutilation phenotype. 735 01:00:32,260 --> 01:00:37,090 They have a mental retardation, a lot of neurological stuff. 736 01:00:37,090 --> 01:00:39,880 And no one can really explain why 737 01:00:39,880 --> 01:00:44,740 it is that one gets this complex neurological phenotype 738 01:00:44,740 --> 01:00:49,570 because you can't salvage guanine and hypoxanthine. 739 01:00:49,570 --> 01:00:53,740 The salvage of purines, which, again, I show here 740 01:00:53,740 --> 01:01:00,220 on a slide, really that shows all of the different enzymes 741 01:01:00,220 --> 01:01:02,980 and ways here is that HPRT to salvage 742 01:01:02,980 --> 01:01:07,570 guanine and hypoxanthine to make IMP and GMP. 743 01:01:07,570 --> 01:01:11,350 There's actually quite complex enzymes 744 01:01:11,350 --> 01:01:15,340 that can allow you to basically interconvert 745 01:01:15,340 --> 01:01:19,330 all of these nucleosides with nucleotides as well 746 01:01:19,330 --> 01:01:23,410 as with the free purine bases. 747 01:01:23,410 --> 01:01:27,910 Each of these enzymes has its own syndrome associated with it 748 01:01:27,910 --> 01:01:30,820 if there's humans that are deficient. 749 01:01:30,820 --> 01:01:33,940 Some of it causes immune system problems. 750 01:01:33,940 --> 01:01:35,860 There's the mental problems I talked about 751 01:01:35,860 --> 01:01:38,170 with HPRT deficiency. 752 01:01:38,170 --> 01:01:42,370 Again, why certain enzymes and pathways 753 01:01:42,370 --> 01:01:46,510 are more important in specific cells and tissues, 754 01:01:46,510 --> 01:01:49,330 not so well understood. 755 01:01:49,330 --> 01:01:51,160 And really, what I want you to appreciate 756 01:01:51,160 --> 01:01:53,020 is that this is complicated and just 757 01:01:53,020 --> 01:01:56,530 realize that this happens in case you encounter it 758 01:01:56,530 --> 01:01:58,360 in your first future career and you 759 01:01:58,360 --> 01:02:01,020 want to come back and study it. 760 01:02:01,020 --> 01:02:01,520 OK. 761 01:02:05,870 --> 01:02:08,780 All right, so we just went through the way 762 01:02:08,780 --> 01:02:12,110 that you can salvage purines or make purines de novo. 763 01:02:12,110 --> 01:02:15,380 And of course, that gives you ribonucleotides that 764 01:02:15,380 --> 01:02:18,260 can be used to generate RNA. 765 01:02:18,260 --> 01:02:20,735 If you're going to generate DNA, you then 766 01:02:20,735 --> 01:02:23,750 need to, instead of generating the ribonucleotides, 767 01:02:23,750 --> 01:02:27,830 end up with the deoxyribonucleoside 768 01:02:27,830 --> 01:02:29,300 triphosphates. 769 01:02:29,300 --> 01:02:30,480 And what does that mean? 770 01:02:30,480 --> 01:02:32,270 So if we draw here, here's a-- 771 01:02:43,280 --> 01:02:48,260 for whatever base, adenine, guanine, whatever, basically, 772 01:02:48,260 --> 01:02:51,920 if we're going to generate a deoxyribonucleoside, 773 01:02:51,920 --> 01:02:55,430 we have to-- remember, this is the 2 position of ribose. 774 01:02:55,430 --> 01:02:59,510 And so we need to reduce this 2 position of ribose-- 775 01:02:59,510 --> 01:03:02,810 basically, reduce it from the alcohol 776 01:03:02,810 --> 01:03:17,160 to the saturated hydrocarbon to generate 777 01:03:17,160 --> 01:03:21,390 the deoxyribonucleoside. 778 01:03:21,390 --> 01:03:28,870 This is carried out by an enzyme called 779 01:03:28,870 --> 01:03:30,820 ribonucleotide reductase. 780 01:03:39,400 --> 01:03:42,330 Makes sense-- we're doing a redox reaction, 781 01:03:42,330 --> 01:03:47,010 reducing that alcohol to saturated hydrocarbon, 782 01:03:47,010 --> 01:03:49,860 ribonucleotide reductase. 783 01:03:49,860 --> 01:03:51,930 Interesting mechanism, you can look it up 784 01:03:51,930 --> 01:03:53,190 if you're interested. 785 01:03:53,190 --> 01:03:56,610 Ribonucleotide reductase, I drew it this way on purpose, 786 01:03:56,610 --> 01:04:04,500 always acts on the dNDP, so the nucleoside diphosphate-- 787 01:04:04,500 --> 01:04:07,680 sorry, on the nucleoside diphosphate 788 01:04:07,680 --> 01:04:12,330 to generate the deoxynucleoside-- 789 01:04:12,330 --> 01:04:15,810 sorry, deoxynucleotide diphosphate. 790 01:04:15,810 --> 01:04:21,060 So the nucleotide diphosphate is what ribonucleotide reductase 791 01:04:21,060 --> 01:04:27,180 acts on to generate the deoxynucleotide diphosphate. 792 01:04:27,180 --> 01:04:33,540 So you would take ADP and generate dADP, GDP, 793 01:04:33,540 --> 01:04:36,830 and generate dGDP. 794 01:04:36,830 --> 01:04:38,630 All right, and we'll come back to this 795 01:04:38,630 --> 01:04:41,000 in a minute for pyrimidines. 796 01:04:41,000 --> 01:04:43,190 Before I get to that, I just want to quickly go 797 01:04:43,190 --> 01:04:45,740 through pyrimidine synthesis. 798 01:04:45,740 --> 01:04:49,970 And so pyrimidine synthesis is a lot simpler 799 01:04:49,970 --> 01:04:52,160 than purine synthesis. 800 01:04:52,160 --> 01:04:55,280 I also like to point out that pyrimidines are also much less 801 01:04:55,280 --> 01:04:57,680 abundant in cells than purines. 802 01:04:57,680 --> 01:05:00,750 So remember, ATP, 10 millimolar concentration in 803 01:05:00,750 --> 01:05:02,960 cells, 1 to 10 millimolar-- 804 01:05:02,960 --> 01:05:07,430 GTP, somewhere in the high hundreds of micromolar. 805 01:05:07,430 --> 01:05:10,190 Purines are more-- sorry, pyrimidines 806 01:05:10,190 --> 01:05:11,990 are more tens of micromolar. 807 01:05:11,990 --> 01:05:16,190 And so orders of magnitude, less pyrimidines in cells than there 808 01:05:16,190 --> 01:05:17,420 are purines. 809 01:05:17,420 --> 01:05:19,580 And that's because GTP, ATP, those 810 01:05:19,580 --> 01:05:22,430 are important energy transduction molecules. 811 01:05:22,430 --> 01:05:24,770 We saw a couple places where pyrimidines can be 812 01:05:24,770 --> 01:05:26,810 energy transduction molecules-- 813 01:05:26,810 --> 01:05:28,800 UTP, glucose, et cetera. 814 01:05:28,800 --> 01:05:33,620 But in general, most reactions use ATP and GTP 815 01:05:33,620 --> 01:05:37,380 as energy sources. 816 01:05:37,380 --> 01:05:41,600 And so whereas pyrimidines are primarily 817 01:05:41,600 --> 01:05:46,310 for building RNA and DNA, and so much less abundant in cells. 818 01:05:46,310 --> 01:05:50,120 All right, so pyrimidine synthesis-- 819 01:05:52,930 --> 01:05:53,625 get my colors. 820 01:05:59,700 --> 01:06:07,190 So pyrimidine synthesis starts with carbamoyl phosphate. 821 01:06:07,190 --> 01:06:10,360 And so you'll remember carbamoyl phosphate 822 01:06:10,360 --> 01:06:16,230 from when we discussed the urea cycle. 823 01:06:16,230 --> 01:06:21,835 OK, so just as a quick reminder, so here's bicarbonate. 824 01:06:21,835 --> 01:06:25,550 I won't use colors to start, just for time. 825 01:06:25,550 --> 01:06:30,130 So if we phosphorylate bicarbonate, 826 01:06:30,130 --> 01:06:36,070 we ended up getting this molecule, that bicarbonate. 827 01:06:36,070 --> 01:06:42,230 We can then have glutamine to glutamate conversion. 828 01:06:42,230 --> 01:06:45,670 That gives us-- releases an ammonia. 829 01:06:45,670 --> 01:06:53,410 That ammonia can release that phosphate. 830 01:06:53,410 --> 01:07:00,610 So ATP to ADP plus phosphate, then gives us 831 01:07:00,610 --> 01:07:08,700 this molecule, which we can phosphorylate again 832 01:07:08,700 --> 01:07:33,520 to give us this molecule, which is carbamoyl phosphate. 833 01:07:33,520 --> 01:07:37,480 Nothing new here, exactly what we showed last time 834 01:07:37,480 --> 01:07:39,940 in the urea cycle. 835 01:07:39,940 --> 01:07:47,470 So carbamoyl phosphate will react with aspartate. 836 01:07:52,840 --> 01:07:58,300 So here's aspartate, the amino acid aspartate. 837 01:08:09,720 --> 01:08:36,479 OK, and that generates this intermediate. 838 01:08:47,350 --> 01:08:50,529 OK, so that generates that intermediate. 839 01:08:50,529 --> 01:08:58,779 This can now undergo ring closure like that. 840 01:09:09,490 --> 01:09:13,649 And then we end up with a molecule 841 01:09:13,649 --> 01:09:45,450 that looks like the familiar ring structure for pyrimidine 842 01:09:45,450 --> 01:09:48,060 shown here, OK? 843 01:09:48,060 --> 01:09:50,160 I drew it this way because the next step, 844 01:09:50,160 --> 01:09:52,669 so this molecule is called dihydroorotate. 845 01:09:58,710 --> 01:10:01,440 And the next step is going to be to oxidize 846 01:10:01,440 --> 01:10:03,990 this carbon-carbon bond. 847 01:10:03,990 --> 01:10:08,610 If we oxidize this carbon-carbon bond, 848 01:10:08,610 --> 01:10:11,520 that gives us hydride ion-- 849 01:10:11,520 --> 01:10:13,620 oxidizing a carbon-carbon bond, just 850 01:10:13,620 --> 01:10:16,770 like we saw in lipid oxidation, just like we 851 01:10:16,770 --> 01:10:19,170 saw in succinate dehydrogenase. 852 01:10:19,170 --> 01:10:24,510 This is an FAD as an electron acceptor. 853 01:10:24,510 --> 01:10:26,910 FAD is reduced to FADH2. 854 01:10:26,910 --> 01:10:29,250 This occurs as an alternative to complex II 855 01:10:29,250 --> 01:10:30,650 in the electron transport chain. 856 01:10:30,650 --> 01:10:33,120 So this would occur in the mitochondrial membrane 857 01:10:33,120 --> 01:10:35,370 and be a way to take dihydroorotate 858 01:10:35,370 --> 01:10:38,250 to orotate, carried out by an enzyme called 859 01:10:38,250 --> 01:10:43,590 dihydroorotate dehydrogenase or DHODH, which ends up 860 01:10:43,590 --> 01:10:47,350 forming this alternate electron transport chain, 861 01:10:47,350 --> 01:10:49,950 electrons ultimately ending up as oxygen. 862 01:10:49,950 --> 01:10:56,050 And this generates the base pyrimidine, the IMP equivalent, 863 01:10:56,050 --> 01:10:58,860 if you will, of a pyrimidine, which 864 01:10:58,860 --> 01:11:31,500 is this molecule, which is this molecule called orotate. 865 01:11:31,500 --> 01:11:40,770 And then orotate will react with PRPP, 866 01:11:40,770 --> 01:11:44,830 generating a pyrophosphate, which 867 01:11:44,830 --> 01:11:47,840 can be cleaved to two inorganic phosphates, 868 01:11:47,840 --> 01:11:49,900 pull the reaction forward. 869 01:11:49,900 --> 01:11:53,770 And we end up with this phosphoribose. 870 01:11:57,030 --> 01:12:03,330 And basically, this is going to add to this nitrogen right 871 01:12:03,330 --> 01:12:04,080 here. 872 01:12:04,080 --> 01:12:05,790 And so you're used to seeing it drawn 873 01:12:05,790 --> 01:12:07,920 if I add to that nitrogen, and now 874 01:12:07,920 --> 01:12:12,360 rotate the molecule in that direction. 875 01:12:36,790 --> 01:12:42,490 And you get this molecule, which is orotate monophosphate, 876 01:12:42,490 --> 01:12:48,400 orotate monophosphate, which is the starting pyrimidine. 877 01:12:48,400 --> 01:12:57,010 And simply decarboxylating orotate monophosphate 878 01:12:57,010 --> 01:13:31,830 will give us the pyrimidine, uridine monophosphate or UMP. 879 01:13:31,830 --> 01:13:39,990 OK, and so UMP generated in that way. 880 01:13:39,990 --> 01:13:46,110 Now to turn UMP into CTP, what happens 881 01:13:46,110 --> 01:13:50,220 is UMP is phosphorylated twice. 882 01:13:50,220 --> 01:13:52,320 So that's going to cost two ADP. 883 01:13:52,320 --> 01:13:55,110 That's going to generate a UTP. 884 01:13:55,110 --> 01:13:57,930 And then we're going to take that UTP, 885 01:13:57,930 --> 01:14:01,650 and carry out, again, the same reaction that you saw earlier 886 01:14:01,650 --> 01:14:07,650 for FGAR to FGAM, xanthine monophosphate to guanine 887 01:14:07,650 --> 01:14:09,420 monophosphate. 888 01:14:09,420 --> 01:14:11,820 So I'm going to take glutamine to glutamate, 889 01:14:11,820 --> 01:14:20,750 transfer the ammonia, take ATP to ADP plus Pi. 890 01:14:20,750 --> 01:14:23,510 And basically what we're going to end up doing 891 01:14:23,510 --> 01:14:30,220 is changing that double bond oxygen, so double bond there, 892 01:14:30,220 --> 01:14:31,990 and an amino group on top. 893 01:14:31,990 --> 01:14:59,110 And that is how we generate the other major purine, 894 01:14:59,110 --> 01:15:01,880 cytidine triphosphate-- or another major pyrimidine, 895 01:15:01,880 --> 01:15:04,040 cytidine triphosphate. 896 01:15:04,040 --> 01:15:10,040 So UTP to CTP reaction that we've already seen before 897 01:15:10,040 --> 01:15:13,220 gets us CTP. 898 01:15:13,220 --> 01:15:18,230 Now to get deoxycytidine, obviously, 899 01:15:18,230 --> 01:15:21,410 remember ribonucleotide reductase acts on 900 01:15:21,410 --> 01:15:25,460 the diphosphate, so release of phosphates. 901 01:15:25,460 --> 01:15:27,740 So now we have cytidine diphosphate. 902 01:15:27,740 --> 01:15:33,020 And now ribonucleotide reductase can generate 903 01:15:33,020 --> 01:15:36,710 deoxycytidine diphosphate. 904 01:15:36,710 --> 01:15:39,320 Now remember, uracil is not used in DNA. 905 01:15:39,320 --> 01:15:41,570 Instead of uracil, we use thymidine. 906 01:15:41,570 --> 01:15:43,640 So there's no deoxyuracil, or you 907 01:15:43,640 --> 01:15:45,980 don't want to make deoxyuracil because that 908 01:15:45,980 --> 01:15:47,720 will incorporate into DNA. 909 01:15:47,720 --> 01:15:49,530 That's undesirable. 910 01:15:49,530 --> 01:15:54,080 And so the way you make thymidine is you take dCDP 911 01:15:54,080 --> 01:15:57,740 and you further remove a phosphate from it. 912 01:15:57,740 --> 01:15:59,025 And now you get dCMP. 913 01:16:02,090 --> 01:16:06,905 And then you take the dCMP, and you convert it back to dUMP. 914 01:16:10,690 --> 01:16:15,370 Converting it back to dUMP is basically just losing 915 01:16:15,370 --> 01:16:17,290 the nitrogen. We've seen that before when 916 01:16:17,290 --> 01:16:19,030 we broke down guanine-- 917 01:16:19,030 --> 01:16:22,180 same reaction, dCMP back to dUMP. 918 01:16:22,180 --> 01:16:29,830 And then dUMP is a substrate for an enzyme called thymidylate 919 01:16:29,830 --> 01:16:40,140 synthase, which will take uracil and add 920 01:16:40,140 --> 01:16:47,750 a one-carbon unit there, which is methyl group as D 921 01:16:47,750 --> 01:16:50,000 and generate thymidine. 922 01:16:50,000 --> 01:16:51,810 So where does that come from? 923 01:16:51,810 --> 01:16:54,500 Well, we're going to add a methyl group. 924 01:16:54,500 --> 01:16:56,270 We could do that from the folate. 925 01:16:56,270 --> 01:16:59,840 Now you might guess that you'd use 5-methyl-THF for this, 926 01:16:59,840 --> 01:17:01,850 but you actually don't. 927 01:17:01,850 --> 01:17:14,080 Instead, what you use is this N5, 928 01:17:14,080 --> 01:17:24,790 N10-methyl methylene THF, which we got directly from serine 929 01:17:24,790 --> 01:17:27,670 to glycine conversion. 930 01:17:27,670 --> 01:17:50,910 And so this then is going to add the methyl group to have dUMP. 931 01:17:54,010 --> 01:17:56,710 So we're going to add a methyl group there. 932 01:18:10,440 --> 01:18:15,250 And this is going to give me dTMP. 933 01:18:15,250 --> 01:18:17,580 This is what thymidylate synthase does. 934 01:18:17,580 --> 01:18:19,410 But I've added a methyl group. 935 01:18:19,410 --> 01:18:23,647 Remember, this is a formaldehyde oxidation state. 936 01:18:23,647 --> 01:18:25,230 So if I'm going to add a methyl group, 937 01:18:25,230 --> 01:18:27,540 that means that one-carbon unit is getting 938 01:18:27,540 --> 01:18:29,940 reduced when it's being added. 939 01:18:29,940 --> 01:18:32,700 If it's being reduced, something else has to be oxidized. 940 01:18:32,700 --> 01:18:37,840 And the thing that gets oxidized is the folate itself. 941 01:18:37,840 --> 01:18:41,670 So this is-- the tetrahydrofolate 942 01:18:41,670 --> 01:18:47,520 is oxidized to instead be a dihydrofolate, 943 01:18:47,520 --> 01:18:50,170 so double bond here. 944 01:18:50,170 --> 01:18:52,950 So that's oxidation. 945 01:18:52,950 --> 01:18:56,444 Now it becomes dihydrofolate. 946 01:19:00,180 --> 01:19:02,550 And that dihydrofolate, of course, 947 01:19:02,550 --> 01:19:09,000 has to be re-reduced back to the tetrahydrofolate 948 01:19:09,000 --> 01:19:11,910 and pick up a one-carbon unit. 949 01:19:11,910 --> 01:19:18,390 And so this comes from NADPH to NADP+. 950 01:19:18,390 --> 01:19:22,680 So oxidze NADPH, reduce dihydrofolate back 951 01:19:22,680 --> 01:19:26,160 to tetrahydrofolate, pick up another one-carbon unit 952 01:19:26,160 --> 01:19:29,550 from serine to glycine conversion. 953 01:19:29,550 --> 01:19:34,770 And in the end, this is how you synthesize thymidine. 954 01:19:34,770 --> 01:19:36,670 I went through this very quickly. 955 01:19:36,670 --> 01:19:38,820 But this is actually a very important reaction 956 01:19:38,820 --> 01:19:40,320 pharmacologically. 957 01:19:40,320 --> 01:19:43,450 This is an important target for antibiotics. 958 01:19:43,450 --> 01:19:47,310 So the antibiotic Bactrim, a very famous antibiotic, 959 01:19:47,310 --> 01:19:55,140 basically blocks this step called dihydrofolate reductase, 960 01:19:55,140 --> 01:20:00,510 dihydrofolate reductase, DHFR, basically taking dihydrofolate 961 01:20:00,510 --> 01:20:04,770 back to tetrahydrofolate before it can pick up 962 01:20:04,770 --> 01:20:06,840 a serine-glycine unit. 963 01:20:06,840 --> 01:20:11,760 Effects on the bacteria-- very effective antibiotic. 964 01:20:11,760 --> 01:20:16,920 Also, antifolate chemotherapies-- methotrexate, 965 01:20:16,920 --> 01:20:19,440 inhibitor of dihydrofolate reductase, 966 01:20:19,440 --> 01:20:22,710 one of the first chemotherapies ever discovered, 967 01:20:22,710 --> 01:20:27,150 also works because it blocks thymidine synthesis 968 01:20:27,150 --> 01:20:30,630 by preventing your ability to recycle the dihydrofolate back 969 01:20:30,630 --> 01:20:35,250 to tetrahydrofolate and carry out thymidine synthesis. 970 01:20:35,250 --> 01:20:39,150 And so ends up being a pharmacologically, 971 01:20:39,150 --> 01:20:42,810 at least for human medicine, very important series 972 01:20:42,810 --> 01:20:45,450 of reactions. 973 01:20:45,450 --> 01:20:47,225 All right, so for pyrimidine salvage, 974 01:20:47,225 --> 01:20:48,850 there's almost nothing to say about it. 975 01:20:48,850 --> 01:20:51,690 You just take the pyrimidine bases, and you add it to PRPP. 976 01:20:51,690 --> 01:20:54,900 And now you have your pyrimidine back. 977 01:20:54,900 --> 01:20:57,900 And so nothing more to say there. 978 01:20:57,900 --> 01:21:01,680 And really, what I want to do is close with a few comments 979 01:21:01,680 --> 01:21:09,150 about regulation of nucleoside and nucleotide metabolism. 980 01:21:09,150 --> 01:21:12,390 And so obviously, if you're going 981 01:21:12,390 --> 01:21:17,190 to synthesize RNA and DNA, you need 982 01:21:17,190 --> 01:21:22,080 to have the right ratios of nucleotides. 983 01:21:22,080 --> 01:21:26,580 So if your RNA polymerase is making an RNA molecule 984 01:21:26,580 --> 01:21:31,350 and it doesn't have the right base, it can't keep going. 985 01:21:31,350 --> 01:21:34,830 And this is particularly a problem for synthesis 986 01:21:34,830 --> 01:21:37,320 because remember, and as you will learn or have learned 987 01:21:37,320 --> 01:21:39,850 in cell biology, you have to license 988 01:21:39,850 --> 01:21:41,910 your origins of replication before you 989 01:21:41,910 --> 01:21:44,440 start DNA replication. 990 01:21:44,440 --> 01:21:47,620 And that prevents you from ever replicating 991 01:21:47,620 --> 01:21:50,080 the same amount of DNA twice, meaning you say, 992 01:21:50,080 --> 01:21:51,100 here's where you start. 993 01:21:51,100 --> 01:21:53,740 And once you start, you can't stop until you 994 01:21:53,740 --> 01:21:55,780 finish DNA replication. 995 01:21:55,780 --> 01:21:58,000 Well, this becomes a tremendous problem 996 01:21:58,000 --> 01:21:59,980 if you're trying to replicate your DNA and you 997 01:21:59,980 --> 01:22:02,920 run out of nucleosides or nucleotides. 998 01:22:02,920 --> 01:22:06,430 And so there has to be complex mechanisms in place 999 01:22:06,430 --> 01:22:09,100 to make sure that cells always have 1000 01:22:09,100 --> 01:22:14,440 the right levels of nucleotide triphosphates 1001 01:22:14,440 --> 01:22:18,970 and deoxynucleotide triphosphates. 1002 01:22:18,970 --> 01:22:22,060 And so there's complex feedbacks that we 1003 01:22:22,060 --> 01:22:24,510 could give a whole lecture on. 1004 01:22:24,510 --> 01:22:26,860 But for the purposes of today, I just 1005 01:22:26,860 --> 01:22:30,250 want you to be familiar and know that they're here. 1006 01:22:30,250 --> 01:22:32,620 And they also all make sense. 1007 01:22:32,620 --> 01:22:36,410 They fit exactly with what we've talked about in other pathways. 1008 01:22:36,410 --> 01:22:39,250 And so shown here on the slide is the control 1009 01:22:39,250 --> 01:22:40,940 of purine synthesis. 1010 01:22:40,940 --> 01:22:42,800 And so this makes perfect sense. 1011 01:22:42,800 --> 01:22:46,000 So if you have plenty of purines, 1012 01:22:46,000 --> 01:22:49,175 meaning you have a lot of AMP, GMP, IMP. 1013 01:22:49,175 --> 01:22:50,050 Well, what do you do? 1014 01:22:50,050 --> 01:22:54,220 Well, don't charge the ribose 5-phosphate to PRPP. 1015 01:22:54,220 --> 01:22:57,640 Or don't carry out the first step in purine synthesis 1016 01:22:57,640 --> 01:23:00,160 where you take that ammonia from glutamine 1017 01:23:00,160 --> 01:23:05,350 and take PRPP and add it to the PRPP to begin purine synthesis. 1018 01:23:05,350 --> 01:23:08,680 And so all of these purine monophosphates 1019 01:23:08,680 --> 01:23:13,570 inhibit this early steps of purine synthesis, makes sense. 1020 01:23:13,570 --> 01:23:17,200 What if you have an imbalance in your AMP and GMP pools? 1021 01:23:17,200 --> 01:23:19,330 Well, the way you prevent that, remember, 1022 01:23:19,330 --> 01:23:22,330 we already discussed that to turn IMP into AMP, 1023 01:23:22,330 --> 01:23:27,460 you use GTP as an energy phosphate donor. 1024 01:23:27,460 --> 01:23:32,450 And to turn IMP into GMP, you use ATP as an energy phosphate 1025 01:23:32,450 --> 01:23:32,950 donor. 1026 01:23:32,950 --> 01:23:35,980 So that's one way you regulate this. 1027 01:23:35,980 --> 01:23:38,560 But the other thing is if you have enough AMP, well, 1028 01:23:38,560 --> 01:23:39,940 don't try to make AMP. 1029 01:23:39,940 --> 01:23:42,880 And if you have enough GMP, don't try to make GMP. 1030 01:23:42,880 --> 01:23:45,070 And so there's feedbacks there as well 1031 01:23:45,070 --> 01:23:49,650 that add to balancing the AMP and GMP pools. 1032 01:23:49,650 --> 01:23:50,850 What about pyrimidines? 1033 01:23:50,850 --> 01:23:56,400 Well, pyrimidines-- simpler synthesis, simpler regulation. 1034 01:23:56,400 --> 01:24:00,210 If you have enough pyrimidines, enough CTP, 1035 01:24:00,210 --> 01:24:02,430 don't do the first step where you 1036 01:24:02,430 --> 01:24:05,100 take aspartate and carbamoyl phosphate 1037 01:24:05,100 --> 01:24:13,480 and generate that first step here in pyrimidine synthesis. 1038 01:24:13,480 --> 01:24:16,600 dNTPs, obviously, as I said, is particularly critical 1039 01:24:16,600 --> 01:24:19,080 because once you start replicating your DNA, 1040 01:24:19,080 --> 01:24:21,100 you always have to have the right balance 1041 01:24:21,100 --> 01:24:23,290 of deoxyribonucleotides. 1042 01:24:23,290 --> 01:24:26,350 And so ribonucleotide reductase, that 1043 01:24:26,350 --> 01:24:31,540 enzyme up there that basically balances and synthesizes 1044 01:24:31,540 --> 01:24:36,520 your deoxyribonucleotides, it has incredibly complex feedback 1045 01:24:36,520 --> 01:24:40,150 mechanisms that one could spend a whole lecture talking about. 1046 01:24:40,150 --> 01:24:43,270 But effectively, it has various feedback mechanisms 1047 01:24:43,270 --> 01:24:46,690 with allosteric sites and other feedbacks that are still, 1048 01:24:46,690 --> 01:24:50,590 honestly, being worked out, that, in the end, 1049 01:24:50,590 --> 01:24:52,720 is a way that helps you balance to make 1050 01:24:52,720 --> 01:24:54,550 sure you have the right levels of all 1051 01:24:54,550 --> 01:24:57,370 the different deoxyribonucleotides 1052 01:24:57,370 --> 01:24:59,260 in the end. 1053 01:24:59,260 --> 01:25:04,480 All right, if you're interested more in the feedbacks 1054 01:25:04,480 --> 01:25:06,520 of ribonucleotide reductase, these 1055 01:25:06,520 --> 01:25:09,440 are covered well in your textbook. 1056 01:25:09,440 --> 01:25:12,400 Any of the topics that we didn't have time 1057 01:25:12,400 --> 01:25:17,080 to cover that might become relevant for your future 1058 01:25:17,080 --> 01:25:22,000 endeavors in biology, medicine, metabolism, whatever, 1059 01:25:22,000 --> 01:25:23,740 all of those are well covered. 1060 01:25:23,740 --> 01:25:28,120 And hopefully, if you could absorb some of the concepts 1061 01:25:28,120 --> 01:25:32,800 that I tried to impart to you about how metabolism works, 1062 01:25:32,800 --> 01:25:36,050 you can understand those pathways better. 1063 01:25:36,050 --> 01:25:39,070 Hope you enjoyed 7.05. 1064 01:25:39,070 --> 01:25:42,640 Hope you enjoyed learning about metabolism. 1065 01:25:42,640 --> 01:25:46,350 And thank you very much for your attention.