Lecture 16. Glycobiology & Polyketides, Part 2.
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| Teil | 16 | |
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ChemieingenieurinEntzündungChemische ForschungMagmaAmalgamHalbedelsteinPolysaccharideKohlenhydratchemieProteineVerstümmelungKalisalzeCalciumhydroxidOmega-3-FettsäurenHydroxybuttersäure <gamma->PolyketideChromosomenkondensationMolekülEukaryontische ZelleTriterpenePolyketideTopizitätBindegewebeGezeitenküsteBiosyntheseButterChemische ReaktionChemische StrukturFeuerKrankengeschichteSäureVerbundwerkstoffGesundheitsstörungProteineSymptomatologieErfrischungsgetränkSatelliten-DNSOberflächenchemieSonnenschutzmittelPolysaccharideChemische VerbindungenTiermodellAzokupplungBaseDNS-SyntheseFremdstoffFunktionelle GruppeGelatineKlinisches ExperimentKrebsforschungMeeresspiegelReaktionsmechanismusSubstitutionsreaktionSüßstoffTellerseparatorKonvertierungWursthülleGlykosylierungIonenbindungKonzentratOmega-6-FettsäurenKondensationsreaktionVerzweigte VerbindungenZuchtzielKaugummiKonformationsisomerieOrlistatBukett <Wein>Energiearmes LebensmittelAlpha-1-RezeptorSucraloseMultiproteinkomplexKohlenhydratchemieChemische BiologieElektronische ZigaretteEntzündungPolymereZuckerFettOligosaccharideOligonucleotideMonosaccharideZunderbeständigkeitHydroxyaldehydeGlykosidasenOrdnungszahlFettsäure-SynthaseChemische ForschungVernetzungsmittelVorlesung/Konferenz
09:26
AcepromazinChemische ForschungAsparaginsäureWasserbeständigkeitLipideGalactosePlasmamembranMedroxyprogesteronEthanolPhosphinEukaryontische ZelleHydrolysatKalisalzeProstaglandineMischenLeukotrien C4EnzymBiosyntheseMolekülBiomembranChemische ReaktionChemische StrukturEntzündungEnzymLipideMolekülRauschgiftSäureVerbundwerkstoffWasserPlasmamembranProteineOrganisches LösungsmittelOberflächenchemieCytoplasmaKonjugateQuerprofilHistidinChemische VerbindungenTiermodellAbschreckenAcetonBindungsenergieCarboxylateEpoxideErdrutschEsterEukaryontische ZelleFließverhaltenFunktionelle GruppeGlutathionPhospholipasenReaktionsmechanismusSerinThiolgruppeVerhungernMakrophageGangart <Erzlagerstätte>Kettenlänge <Makromolekül>AntiasthmatikumEicosanoideMassendichteExtrazellulärraumAsparaginsäureSetzen <Verfahrenstechnik>LactitolIonenbindungTandem-ReaktionOmega-6-FettsäurenDurchflussHydrolysatAktives ZentrumDuplikationFülle <Speise>Domäne <Biochemie>ThioesterChemische ForschungGlättung <Oberflächenbehandlung>Leukotrien C4FeuerKatalysatorKrankengeschichteOsmoseLeukozytLöslichkeitStimulansAktivierung <Chemie>BaseGasphasePasteCarboxylierungWasserbeständigkeitPulverWursthülleTransformation <Genetik>Single electron transferMolekülbibliothekBukett <Wein>Aktivität <Konzentration>AstheniaComputeranimation
18:20
EnzymLeukotrien C4StoffwechselwegProstaglandineAtomChemische ForschungMeerMagmaMolkeAmalgamSchälgangOrnithinGalactoseNatriumhydroxidGlycerinHydrolysatSeifenherstellungPolyketideAcetylsalicylsäureBiomembranChemische ReaktionChemische StrukturEntzündungEnzymFieberInhibitorLipideMolekülSäureSchmerzSyndromHydroxideProteineSatelliten-DNSEthanolProstaglandinePharmazeutische ChemieChemische VerbindungenTiermodellAllmendeAtombindungChemischer ProzessCobaltoxideDoppelbindungErdrutschEukaryontische ZelleGezeitenstromGlycerinHydrophobe WechselwirkungHydroxylgruppeNatriumhydroxidQuelle <Hydrologie>ReaktionsmechanismusSenseSymptomLokalantibiotikumMakrophageGangart <Erzlagerstätte>Kettenlänge <Makromolekül>Posttranslationale ÄnderungAchatSetzen <Verfahrenstechnik>StoffwechselwegWursthüllePentapeptidePräkursorTandem-ReaktionOmega-6-FettsäurenSystemische Therapie <Pharmakologie>StromschnelleAktives ZentrumFülle <Speise>MolekülbibliothekSeifenherstellungBukett <Wein>Beta-FaltblattReglersubstanzBiodieselElektronische ZigarettePlasmamembranFettTankArachidonsäureEnoleEsterProstaglandinsynthaseRöstenSerinVerseifungWasserbeständigkeitPolyketideFettsäure-SynthaseThioesterAktivität <Konzentration>AntigenitätComputeranimation
27:14
AcepromazinChemische ForschungAscheHydroxyaldehydeChromosomenkondensationSäureBiosyntheseChemische VerbindungenPolyketideBukett <Wein>GalactoseStreptomycinMergelImmunglobulin GKalisalzeAssemblyRoher SchinkenZinnerzHalbedelsteinBiskalcitratumBiosyntheseChemische ReaktionEnzymMakromolekülMolekülOrganische ChemiePharmaziePlasmamembranProteineTetracyclineKomplikationOberflächenchemieRibosomKohlenstofffaserGenKetoneChemische VerbindungenAcetonitrilAcylgruppeAromatizitätAzokupplungChemischer ProzessDNS-SyntheseErythromycinEukaryontische ZelleGezeitenstromHydrocarboxylierungIonenpumpeKatalaseMeeresspiegelPropionsäureReaktionsmechanismusReduktionsmittelThermoformenBenzolringGangart <Erzlagerstätte>Kettenlänge <Makromolekül>OperonPulverAlkoholische GärungProteinsynthesePolyketideSpanbarkeitSekundärstrukturAmphotericin BIonenbindungPräkursorClaisen-KondensationFettsäure-SynthaseAldolreaktionAktives ZentrumFülle <Speise>Offener LeserahmenFreies ElektronChromosomenkondensationDomäne <Biochemie>MultiproteinkomplexChemische ForschungAntibiotikaresistenzSäureProtein CPhasengleichgewichtSubstrat <Boden>TiermodellElektronentransferNeutralisation <Chemie>ZunderbeständigkeitLokalantibiotikumGezeitenküsteWursthülleBoyle-Mariotte-GesetzOmega-6-FettsäurenAtomsondeSingle electron transferTopizitätVorlesung/Konferenz
36:07
PolyketideChemische ForschungAssemblyAcepromazinSynthasenQuellgebietHeterodimereGalactoseAmalgamCalciumhydroxidChemische StrukturEnzymGummiMolekülMolekularbiologieOxidschichtSchmerzStahlWasserHydroxideProteineChemieanlageKohlenstofffaserSynthesekautschukQuerprofilTiermodellBaseElektronentransferErdrutschEukaryontische ZelleFunktionelle GruppeGezeitenstromHydroxylgruppeReaktionsmechanismusReduktionsmittelSenseThermoformenTriterpeneGangart <Erzlagerstätte>LebkuchenAbbruchreaktionSetzen <Verfahrenstechnik>Infrastruktur <Histologie>StereoselektivitätWursthüllePentapeptideSpanbarkeitSekundärstrukturIonenbindungPräkursorOmega-6-FettsäurenSingulettzustandKondensationsreaktionHydrolysatEnhancerAktives ZentrumHalbedelsteinAssemblySilencerFreies ElektronAlkohole <tertiär->StockfischBukett <Wein>Domäne <Biochemie>HeterodimereMultiproteinkomplexChemische ForschungBiochemikerinElektronische ZigaretteChemische ReaktionKetoneStereochemieDoppelhelixAcylgruppeCytochrom P-450ErythromycinHydrocarboxylierungIsoprenSynthasenPolyketideClaisen-KondensationFettsäure-SynthaseQuellgebietThioesterTafelbild
45:01
Chemische ForschungBiosyntheseHexamereArzneimittelChemische VerbindungenTriterpeneTerpeneMagmaPolypropylenChemische BindungChemische ReaktionHydroxyaldehydeAlkeneBiosyntheseChemische ReaktionChemische StrukturEnzymFormaldehydGeochemieGummiMolekülPolymereSäureWasserZusatzstoffKohlenwasserstoffeEliminierungsreaktionKohlenstofffaserInselMagnesiumPhasengleichgewichtKomplexePhosphateChemische VerbindungenAbschreckenAdditionsverbindungenBaseCobaltoxideDeformationsverhaltenErdrutschGeröllIsomerManganerzMeeresspiegelMolekulardynamikOrangensaftReaktionsgleichungSenseTriterpeneGangart <Erzlagerstätte>Pegel <Hydrologie>WursthülleUmamiPentapeptideSekundärstrukturChemische FormelPräkursorOmega-6-Fettsäurenf-ElementLipogeneseAktives ZentrumOktanzahlChemische EigenschaftMentholAlkohole <tertiär->PyrogallolBukett <Wein>ChromosomenkondensationQuellgebietTopizitätBeta-FaltblattEisflächeReglersubstanzMultiproteinkomplexChemische ForschungMalerfarbeClaisen-UmlagerungGlutaminsäureStereochemieEthanCyclische VerbindungenAdenylatcyclaseCarbanionCarbokationDipeptideEukaryontische ZelleFunktionelle GruppeMesomerieTerpeneThermoformenIsoprenKettenlänge <Makromolekül>SynthasenAllylverbindungenPolyketideHydroxyaldehydeIonenbindungDiphosphateTaxolIsopentenylpyrophosphatPrenylgruppeGeraniolElektronische ZigaretteVorlesung/Konferenz
53:55
Chemische ForschungChemische ReaktionCholesterinEnzyminhibitorHydroglimmerPhosphateDiphosphateKettenlänge <Makromolekül>TerpenePolypeptidketten bindende ProteineTonmineralBiosyntheseChemische ReaktionCholesterinEnzymMolekülSäureWasserProteineZusatzstoffRückstandEliminierungsreaktionKohlenstofffaserMagnesiumChemische VerbindungenCyclische VerbindungenQuarz <alpha->AlterungAzokupplungBindungsenergieCarbokationCarboxylateDecarboxylierungEnoleErdrutschEsterMündungFunktionelle GruppeLactoneLipidsenkerPhosphorylierungReaktionsmechanismusReduktionsmittelThermoformenTiefseeÜbergangszustandIsoprenGangart <Erzlagerstätte>Kettenlänge <Makromolekül>SynthasenSetzen <Verfahrenstechnik>MevalonsäureIonenbindungPräkursorDiphosphateAldolreaktionHydrolysatSeitenketteAktives ZentrumLagerungFülle <Speise>Alkohole <tertiär->KarsthöhleAktivität <Konzentration>PrenylgruppeFleischersatzEisenFormaldehydKohleKomplikationGenAromatizitätEnzyminhibitorHydroxymethylglutaryl-CoA-ReductaseKatalaseKrebsforschungMeeresspiegelPipetteSchussverletzungSenseCarboxylierungWursthülleSekundärstrukturTechnikumsanlageMannoseOktanzahlAcetyl-CoAPotenz <Homöopathie>HMG-ProteineChemische ForschungComputeranimationTafelbild
01:02:48
Vorlesung/Konferenz
Transkript: Englisch(automatisch erzeugt)
00:07
Okay, we're back. Today we're going to be talking about polyketides and then we'll be on to terpenes. So actually I expect to cover a little bit of terpenes today and then finish off the discussion on Tuesday.
00:22
And then on Tuesday we're on to Chapter 9 to discuss cell signaling and small molecules and cell signaling. And actually I'm going to touch on that topic today. Okay, last time we saw that oligosaccharides are quite heterogeneous and quite diverse.
00:41
That there is no one oligosaccharide component per protein that's encoded by DNA in some way. Rather we saw there was a sort of a random distribution of different structures really on the surface of the cell. And this randomness is a key attribute
01:01
that affects their function, right? We discussed for example how oligosaccharides cause proteins to fold and so having a, you might imagine that having a diversity of different structures might also give you a range of different confirmations accessed by the proteins. Now that latter remark is a little bit speculative
01:20
but that's one of the ways that people are thinking about these things. Okay, so we also discussed how sulfated polysaccharides avidly take up water and if you were blowing your nose on the way into work this morning or the way into class this morning you definitely know about that. We talked about mucus and mucins. We talked a little bit about how O-linked glycans are added one at a time.
01:43
Actually this is good. This is making a couple of points that I meant to make but might have lost over. So the O-linked glycans are added one monosaccharide at a time to build up to big structures. And the N-linked glycans are added as complex branched polysaccharides sometimes in segments of three which we saw
02:01
and sometimes even more complex. After the synthesis of these N and O-linked glycans there's lots of changes that take place through trimming by glycosidases. These are the little scissors that come along and snip apart glycosidic bonds and they do it in sort of semi-random fashion
02:21
and that introduces additional diversity that further complicates our lives if we want to study these things. We also talked about how these oligo polysaccharides are exclusively found on the surface of the cell and are very heterogeneous in composition. So there are really no glycosylated proteins found
02:41
inside the cell although I'm looking for someone to prove me wrong on that one. I know there'll be someone in this class or someone watching this video and that would be great. For now our dogma is that there are no polysaccharides that are attached to proteins found inside the cell. Rather this is sort of an exterior attribute that's grafted on to proteins found on the surface, on the outside
03:02
of the cell and we describe this as a gumball model for the cell where the outside of the cell is sweet with carbohydrates and then the inside is chock full of proteins and DNA where gumballs of course have gelatin as the protein.
03:21
Then we discussed how too much glucose, too many gumballs leads to protein cross-linking and inflammation and how this leads to diseases like diabetes, et cetera and how these are really unavoidable consequences of a western diet that we eat such high concentrations
03:41
of carbohydrates that it's kind of inevitable that we end up with some inflammation resulting from this sort of thing and so this is one of the major challenges I think going forward is now to apply this knowledge about chemical biology, about the underlying mechanisms that are taking place at the level of atoms and bonds
04:01
and try to convince other people in the country to change their lifestyle really. That's a non-trivial challenge, that challenge of the sort of next era which is going from atoms and bonds and then going to society. That's really a major link that I leave to you to take up.
04:22
All right, were there any questions about what we saw last time about carbohydrates, about glycosides, anything you want to know about carbohydrates I will endeavor to answer. I don't know everything but I'll do my best. Stump the chump, yep, yeah, okay, so, okay,
04:53
so the question, let me just repeat the question in case you didn't hear it. The question was you discussed those sugar substitutes
05:00
and aren't some of those sugar substitutes linked to other diseases. Okay, so I'll answer your question in a moment but before I do, I think you've raised a really good point. The sugar substitutes that I presented towards the end, compounds like the thing isolated from stevia, sucralose, et cetera, those aren't a panacea.
05:21
They are not completely benign and one of the astounding things that we find is that those sugar substitutes, even though they're not digested, even though they don't directly produce calories, do seem to result in caloric intake by animals who are fed the sugar substitute. And our theory, our best guess of what's going on is that there's actually sweet receptors not just
05:42
on your tongue but also that are in your stomach that are kind of gauging what kind of food is present. And when those sweet receptors get stimulated by sugar substitutes, they sort of actively start taking up a lot more calories than they otherwise would. And so, you know, even though your soft drink might be zero
06:01
calories, it has the fact of stimulating you to take up more calories than you'd otherwise take up because your body function is expecting something sweet and sugary. Okay, so that's one aspect of what's going on. You know, there's just no free lunch basically. Okay, now the second aspect is the linkage to cancer and I'll be honest, I don't know very much about that,
06:23
about the, you know, the latest research on that sort of thing. I think there was, there's kind of a history looking at certain sweeteners in that context. And some of those kind of early, I'm talking about like 1970s kind of trials were not done
06:41
with the kind of rigor that we would use today. So I think it's time to go back and reevaluate our thinking about that sort of thing using different standards that we now apply. So for example, a lot of those involve really high concentrations of sugar substitutes where it's just physiologically unrealistic. Although I don't know. So people drink liters upon liters of tight soft drink.
07:03
So maybe that is actually kind of a list. So anyway, someone who can design better experiments needs to go back and look at it more closely. And that's all I have to say about that. But thanks for asking. Other questions? Anything you want to know about carbohydrates, monosaccharides,
07:20
polysaccharides? Okay, in the back. Oh, okay, thanks for asking. That's a great question. And I actually don't know your name. Ty. Okay, so Ty's question is how fast do the monosaccharides get added? And the answer is really fast. Like these are being added on the sub-second time scale.
07:44
So on the order of like milliseconds. These reactions are happening very quickly. They're nicely enzyme catalyzed and the reactions are not slow. These are really fast reactions. So, and I think your question would be a really interesting one
08:01
to compare against say DNA being added during DNA polymerization. And I don't have the exact numbers for that. And I think that's a great food for thought question. So thanks for asking. Other questions? All right, let's push on. When we left off last time, we were talking about polysaccharides.
08:20
Or sorry, not polysaccharides, terpenes. And just give me a moment to find where we left off. And then we'll start again. One moment. All right, so I showed you the fatty acid synthase as kind
08:41
of our final thought. And I think maybe what I'll do is I'll start there, OK? So we saw that the fatty acid synthase was this really complex machine that allows the synthesis of fats. And it does this by having this robot arm that cycles around, OK?
09:00
And we saw a key reaction in here which was this clazing condensation reaction where it's basically an aldol that involves a thioester. OK, now take a look at this reaction really closely, OK? I want to give a quiz about this and I want you to be able
09:21
to redo this reaction for me, OK? So please look at this. All right, moving on. At the very last step, I kind of glossed over this reaction but it's a really interesting one and I want to take a moment to appreciate this. So we discussed how the robot arm
09:40
that moved the intermediates amongst the various enzyme domains was linked to the intermediates through a thioester. And that was that phosphopentathienyl arm that I showed you. And so the very last step is this thioesterase that has to hydrolyze this thioester.
10:01
And it's really absolutely notable and it's absolutely fascinating and a great place of convergent evolution or perhaps evolutionary borrowing that the mechanism for this hydrolysis borrows very heavily from the serine proteases down to the point where it has an active site serine, an active site histidine
10:22
and an active site aspartic, aspartic acid. And that's really notable and I think really fun because in the end, that gives us a very rapid way of hydrolyzing these thioesters over here all the way down to a carboxylic acid. And what's great is that we don't really have
10:40
to learn some new mechanism. This is just something that's totally familiar to us. OK. So I want to talk to you about what happens next. So I've shown you how these things are synthesized. I want to talk to you next about how they're applied in the cell and how they're applied in cell signaling. So let's start with what they're used for primarily so these fatty acids form the barrier
11:03
on the cell surface. They're what's composing the plasma membrane. And I want you to recall that the plasma membrane is this layer on that divides, the exterior cellular, extracellular milieu up here from the cytoplasm down here.
11:20
And this barrier is pretty impermeable. It actually, it's really super hydrophobic over here. There are some hydrophilic functionalities that interface between the water and the hydrophobic stuff over here. And I'll look at the, we'll look at the structure in a moment. But the fact here is that the cell very tightly controls what's going to be allowed to pass through its, into its interior,
11:44
into its cytoplasm with one major exception. That major exception is of course pharmaceuticals, right? So small molecule drugs that are hydrophobic are going to be readily able to slip through this semi impermeable barrier, this otherwise impermeable barrier.
12:02
Okay, so all of these compounds in here, all of these chains in here are fatty acids. Let's zoom in and take a closer look. Okay, so when we look at the composition of the plasma membranes found in human cells, what we find is, no surprise, chemical heterogeneity. Okay, we find diversity.
12:20
We find that there are a number of different molecules and I can just give you some percentages of those molecules over here. Okay, so these are average percentages over here and you can see that the plasma membrane is composed of a diverse array of these different, of these different molecules, of these different lipids, okay?
12:42
And the effect here is different changes in the viscosity of the plasma membrane and we find that there are areas of the plasma membrane that are a good deal less viscous than others, that are less fluid than others, that are more viscous than other areas. These are regions that are sometimes called lipid rafts
13:03
where there's even areas where the lipids themselves are insoluble in solvents like acetone. You know, you could take up the cell and usually all this stuff goes neatly into organic solvents like acetone. There's some regions that refuse that kind of treatment. Okay, so anyway, notice that these are ranges
13:21
of different percentages and they range pretty broadly and those ranges have important consequences in terms of the fluidity of the membrane, okay? And the model that you might have learned back in high school, the fluid mosaic model of the cell is true to a certain extent but what we're finding is that it's less fluid,
13:41
that there's actually regions of real density on the cell surface. Okay, now, here's the thing. In order to change, to signal things inside the cell, in order to affect cell function, there's a series of enzymes called phospholipases that play a key role in the cell by hydrolyzing the lipids
14:02
that I showed on the previous slide. So they go through and they hydrolyze these esters or these phosphodiesters in particular spaces where in particular places where each enzyme has a particular specificity for a specific ester, for example. And the consequences here are really fascinating.
14:22
This has the fact of altering cell function by providing molecules that can then bind to proteins and also by altering to a local extent the composition of the membranes that are close to membrane bound proteins.
14:40
Okay, so that's, so we're starting to see that lipids play important roles in cell signaling and I also want to talk to you about cell to cell signaling. Okay, so on this slide over here, this is an example of lipids playing a role of the cells signaling with inside the cell and now I want to talk to you
15:02
about how cells talk to each other. Okay, so there are many different modes of cell-cell communication. Many of them involve the braille communication, the touch, the cell to cell contact that we discussed earlier in this class. But there are also ways for cells to secrete lipid-based molecules that then go off
15:21
and signal other cells to get in on the activity, okay, to get in on the action. And let me show you an example of this. So platelet cells release this TXA2, this should be a subscript, that's released by platelets and that amplifies clot formation. So TXA2 is synthesized
15:40
from a compound called the raccidonic acid. I'll show you the structure of raccidonic acid in a moment. Long story short, you get this compound here that is derived from fatty acids and this has the effect of signaling other platelets to start clotting, to initiate clotting and also to initiate,
16:02
to get smooth muscle to relax in the area to encourage blood flow into the region. Okay, so small molecules are providing important signaling molecules for cell function. It turns out this is actually very complex, okay, and I'm going to just gloss over it. I encourage you to read more about it in the book.
16:22
It's an absolutely fascinating topic and in fact, I can devote a whole lecture to nothing but this. This sort of cell to cell communication involving these leukotrienes and eicosanoids is actually remarkable, okay. So what I'm showing you here are a series
16:43
of different compounds that are synthesized by various enzymes shown in blue. So each one of these blue arrows is an enzyme catalyzed transformation and we'll take a look just at one example of this. So for example, this leukotriene A4 has an epoxide.
17:00
An epoxide of course is a terrific electrophile and in concert with glutathione, this epoxide can be nucleophilically attacked by the thiol from the glutathione to give you this conjugate that now has a thioether bond. This thioether, this compound here keeps going
17:23
until it gets this leukotriene D4 which is then used to stimulate leukocytes, neuronal cells, et cetera and most importantly, bronchial and vascular smooth muscle in the lung. And so, what we're seeing, what I'm trying to show you is a cell signaling cascade
17:40
that eventually leads to inflammation in the lung and asthmatic response. This is what happens when you feel your lungs tightening up as you can't breathe anymore when you're getting an allergic attack or an asthmatic attack. What's happening is all of these signaling molecules are implicated and so this coordinates by having lots
18:03
of signaling molecules running around, this coordinates the response of very diverse cell types ranging from neurons to macrophages and it's absolutely essential that you coordinate these things, right? You wouldn't want for example the bronchial cells contracting and you know constricting unless you had other cells
18:23
that can, you know, redouble efforts to try to deal with whatever it is that's causing their pain, right? You want the macrophages to be an action to chew apart and recognize whatever it is that's causing that, you know, those cells to contract.
18:41
Okay, so there are many different classes of these psychosanoids. They all share a very common core, this arachidonic acid over here and note that this is simply a fatty acid. This can be synthesized using exactly the polyketide synthase that I showed earlier, the fatty acid synthase
19:02
that I showed earlier, the one with the robot arm that moves back and forth between the different subdomains. Okay, let's take a closer look at arachidonic acid. So the first step in the modification of arachidonic acid is absolutely fascinating. There's a class of enzymes called cyclooxygenase
19:23
that add oxygen across this, these two double bonds to give you this prostaglandin molecule. Okay, and this is the very first step in all of the cascades that I've been discussing with you. The very first step here is this oxygen being added by,
19:41
catalyzed by two different enzymes, cyclooxygenase 1 and COX 2. They're abbreviated COX or COX. And all of these, so once this thing is synthesized, there's a whole bunch of other enzymes that then lead to inflammation. Okay, so pathways like these go down to lots
20:01
of different cell types down here and start stimulating inflammation. So if we can block this step up here, then we have a very effective way of dealing with inflammation. And by the way, by inflammation I mean fever, I mean, you know, swelling, I mean like, you know, the immune system starting to spiral out of control.
20:22
Okay, just, you know, walloping on whatever's nearby, including itself. Okay, and that's a bad thing. So for over a century, humans have been using aspirin as a way of combating this pathway. Aspirin, it turns out, is a covalent inhibitor
20:40
of cyclooxygenase 1 and 2. It's fairly nonspecific. And if we zoom in on the cyclooxygenase active site, here is the active site over here. This is arachidonic acid bound to the active site. And in this red stick model right here is a serine hydroxyl that's found in the active site.
21:02
This serine hydroxyl is acetylated by aspirin. So the aspirin transacetylates. It transfers the ester from the aspirin ester to the cyclooxygenase ester from this side to this side. You can imagine this is a very easy reaction, very facile,
21:20
very low energy, analogous to all the transesterase reactions that we saw on Tuesday, right? On Tuesday, I was showing you all the thioester exchange reactions, right? This is totally analogous to that. You eat the aspirin, the aspirin readily flows through your whole body in literally minutes, flowing readily through your cell membranes
21:41
because it's hydrophobic enough. And then when it reaches this active site, it starts attacking this hydroxyl and covalently acetylates it. That has the effect of shutting down this pathway over here, and doing so prevents the cells from responding by inflammation symptoms, okay?
22:04
So that shuts down very quickly the fever or whatever it is that you're dealing with, okay? Make sense? Headaches, fever, et cetera, go on. All right, so this notion that aspirin is actually a covalent inhibitor is a
22:23
relatively new one. I think it was discovered maybe two decades ago or something like that. And so, oh, actually, sorry, the molecular mechanism underlying its covalence, relatively new and absolutely fascinating. Okay, it's actually, I'm pointing this out because it's actually very hard to get molecules
22:40
that covalently modify targets in the cell approved by the Food and Drug Administration. Medicinal chemists in general are leery of compounds that form covalent bonds with targets. And the reason is some misplaced fear of allergenicity, of antigenicity. There's a worry that you're going to modify self-proteins
23:02
and that in turn will stimulate the immune response. And to those who say that, I always point to aspirin as a prime example of why that's not such a big concern. Now, having said that, there is a small percentage of the population that has something, I think, called Rett syndrome. There must be one person in here. We have about a hundred people in this class.
23:21
Is there anyone who's allergic to aspirin? All right, it would be about 1% of the population. So, those people who are allergic to aspirin actually have, you know, stimulated immune response by this, to this covalent addict or perhaps another covalent addict. Okay, so lipids and fats, of course, play a key role
23:44
in our everyday lives as soaps. So, all the lipids and fats that I showed you on the plasma membrane slide can be hydrolyzed quite readily using catalytic amounts of sodium hydroxide. This is typically ly, L-Y-E.
24:03
And this saponification reaction is something that humans have been doing for millennia, really. Most notably, of course, by the protagonist of Fight Club, pro slash anti antagonist of Fight Club who used excess lipids from liposuction
24:22
to make really high-end soaps for boutiques. I happen to love that movie. If you see the movie, check out the microscopy at the very, in the opening credits. It's pretty awesome. If you haven't seen this movie, you should see it just for the saponification scene alone. Okay, here's an old-timey picture showing, you know,
24:42
how historically this process took place. So, when you do this, you can isolate out, depending on how you do it. If you do this in the presence of ethanol, you can isolate out esters, fatty acid esters that basically are nothing more than biodiesel. This is stuff that you can put in the tank of your biodiesel Mercedes Benz and use it
25:02
to drive around to campus. The other thing that's isolated from this is glycerin, which is a very valuable commodity item. It's used pretty extensively as moisturizing soap. And actually, the glycerin soaps are slightly better than the fatty acid soaps. So, when you hydrolyze this with sodium hydroxide
25:22
and no ethanol, you end up with fatty acids. So, you end up with two kinds of soap really. And the glycerin soaps wash away a little bit better than the fatty acid soaps and are slightly more moisturizing. All right, let's see. I think that's all I have to say about fats and lipids.
25:40
And if you have any questions about fats or lipids, I'll take them now. Anything you ever wanted to know about fat but were afraid to ask? All right, what about cellulite? Inevitable consequence of being human. Don't panic. All right, diversification. So, I want to switch gears now and I want to talk to you
26:00
about other kinds of polyketides. I've been showing you the simple ones, OK? But it turns out, this is an amazingly rich class of compounds. This class of molecules extends to all kinds of different antibiotics and all kinds of different circumstances.
26:20
And some of these have nice bitter flavors like this hop constituent. Others, you know, we just don't even know really what they do. But we could find all kinds of examples. So, one way of diversifying these is to have hydroxides in the, you know, these sort of these enolates
26:41
in the structure, these enols in the structure. And that sets you up very neatly for lactonization, this molecule here. So, this is the way that we can start with straight chain precursors and get to rings, OK? This is a very simple way of getting to a ring, right?
27:00
We have an electrophile which is the thioester up here. We have a nucleophile which is this hydroxide. Note that this, even though this is an enol, it's not attacking like an enol. It's actually attack, it's going to form an enol ester. And note too, the compound that results, OK, so this sets you up very neatly for the attack
27:20
because this is perfectly poised in terms of distances of nucleophile and electrophile up here, OK? Beautiful chemistry. So, this is going to set us up to make really complex things. OK, you're ready for complexity? Brace yourself. Here's what can happen. What can happen is you can get to these aromatic compounds
27:43
that I alluded to at the very beginning of this discussion of polyketides. And all you do is you simply start with the straight chain precursors that have carbonyls on them. Recall that this is one of the products from the sequence that I showed when we talked about fatty acid synthases.
28:03
And I showed that it was very quickly, this intermediate was very quickly reduced by NADPH. But here, it doesn't go on to immediate reduction. Instead, this ketone hangs around and it sets you up for an aldol condensation. My all-time favorite reaction, the beautiful aldol,
28:23
object of wonder because our reaction of wonder because it does such an effective job at making carbon-carbon bonds. OK, so this is a really classic reaction. A major challenge, however, is these polyketone things are ridiculously reactive. I mean, you can't really readily isolate them
28:42
so well in the lab. You could freeze them in benzene and work with them. And actually, I have a colleague who does that. They're really nontrivial to work with. The polyketones are so desperate to do this aldol reaction. Everything is so nicely set up because it's going to form this beautiful six-membered ring in intramolecular fashion
29:00
that these are far too reactive to really work with. And it's remarkable that the cell has evolved mechanisms to work with these intermediates. So, a key attribute of this class of enzymes is that they can't have these intermediates kind of bopping around the cell and looking for the next active site. Rather, everything has to be tightly held in place.
29:22
The intermediate has to move from one spot to the next spot to the next spot. And it can't have time to flap around and start making random things. OK. So, here's what can result. And this is the thing that really blows me away. The number of molecules that result is really astonishing
29:42
in its diversity, in its complexity, and its sheer chemical beauty. These things are really wondrous molecules. Tetracycline is a very effective antibiotic. I believe we talked about it that in the context of ribosomal protein synthesis. These, amphotericin is another effective antibiotic.
30:03
This one forms little pores in the cell membrane. And in fact, there's greater than 50 of the 2,000 or so approved pharmaceuticals are directly polyketide and probably another couple hundred or so are derived from polyketides. So, these are synthesized on massive scale,
30:23
not by organic synthesis, rather they are isolated through fermentation by microorganisms. And the microorganisms are like little factories that whip these things out. And they're incredibly valuable, right? Something like tetracycline is sold probably in the ton level per year.
30:41
And it's used quite extensively. OK. And I've even, in the little italics, that actually tells you what organism this can be isolated from. So, each one of these compounds is synthesized by a different microorganism. Now, you can imagine it takes the microorganism a lot of time and energy to make these compounds.
31:02
So, why do you think it is that the microorganism is doing this? Why would a microorganism want to kill some, why would the microorganism want to synthesize compounds that are going to kill microorganisms?
31:21
What's that? Ah, very good. So, it's the competition for scarce resources. These microorganisms are fighting chemical warfare against each other. And they're in this constant arms race to try to develop better chemical compounds that will kill off their competitors.
31:42
And if they succeed, they wipe out all of their competitors and have the field free to themselves to chew on all the tasty stuff that's around. The problem though is that their competitors are also developing ways to nullify and neutralize these compounds.
32:01
And so, these competitors are really, really good. And I think we've seen one example of this. Tetracycline can be readily pumped out of the cell. And so, the competitor microorganisms develop these big pumps that sit at the cell surface and burn ATP and operate night and day. To grab on to tetracycline and other molecules like this
32:23
and simply expel those molecules from the inside up to the outside of the cell. And there's other strategies that we've seen earlier in terms of antibiotic resistance. But it's a constant cat and mouse game between the microorganism that's trying to compete and the other microorganisms
32:40
that are trying not to get killed. So, we chemists are spectators watching from the sidelines of this ongoing multimillion year war. This is an endless war. It's a war that's been going on longer than humans have been on this planet. And this fascinates us. And it gives us all kinds of opportunities to control the pestilent bugs that inflict us as well.
33:04
And so, we do things like grow up these things in large quantities and then use them to kill off the bacteria that are affecting us. Yet at the same time, of course, we're knowing full well that we're highly dependent upon other bacteria that have a symbiotic relationship with us
33:20
and are affected by these compounds equally. All right, I'm a little off topic. I love this topic. So, forgive me. I want to get back to what we want to talk about today. I want to talk to you about the synthesis of complex molecules like this. And good news, a lot of what we've already seen in terms of fatty acid synthase applies
33:42
to much more complicated polyketide synthases. And so, let's talk about how erythromycin is synthesized. So, erythromycin is a very common antibiotic. It's used, I believe, sometimes for treating acne. You might have encountered it at some point. It's used for all kinds of things. It's a pretty effective antibiotic.
34:02
How is it synthesized? It's synthesized in large, it's synthesized by three large macromolecular machines, OK? And these machines are organized in the order of action along the assembly line for this molecule, OK?
34:22
So, again, there's three open reading frames where an open reading frame refers to a single gene that's encoded at the DNA level, translated, or sorry, transcribed into mRNA and then translated as one contiguous protein.
34:41
So, you have three of these that are lined up. Each one of these letters tells us about the identity of a different active site domain. So, a different enzyme active site domain up here, OK? And so, a lot of these are familiar to us. This is the acyl carrier protein, ACP.
35:00
This is the, which attaches the, in this case, propionyl starting material to a thioester, the phosphopentathionyl robot arm. And then, the next step is a ketosynthase. So, that does a Claisen condensation. Next step over here is an acyltransferase.
35:22
That's going to then transfer the intermediate to some other file, a ketoreductase. We've seen ketoreductase. Another acyltransferase, then this gets transferred over to the acyl carrier protein. And then again, another Claisen, another acyltransferase, a ketoreductase, et cetera.
35:41
And so, the product from each of these. So, over here, here's the ketosynthase, here's the product of the Claisen, OK? Oh, after the reduction step takes place. Next step, another ketosynthase, et cetera, to the acyl carrier protein. And so, each one of these ketosynthases elongates a thing
36:01
by three carbons. It introduces a new propionyl fragment. And then, each ketone reductase gives us a new hydroxyl. And note, as usual, the stereochemistry of that hydroxyl is rigidly controlled, OK? And long story short, at the very end over here,
36:21
the last step is carried out by this thioesterase that's actually, rather than doing a conventional hydrolysis of the thioester, it has the hydroxide, the terminal hydroxide come looping in and winging over here to attack this carbonyl, this electrophilic carbonyl of thioester.
36:41
And you can imagine this active site bending the intermediate structure to position this hydroxide right up close to this electrophilic carbonyl. Beautiful chemistry, I'm a big fan. After this is synthesized by this, these polyketide synthases,
37:01
things are passed off to a series of tailoring enzymes that then get in on the action. These include P450, erythra, ARIE F. This is similar to cytochrome P450 that we talked about earlier in the class. This is an oxidase, an effective enzyme for oxidizing things.
37:20
And in this case, it's going to be oxidizing this tertiary carbon right here to give us a tertiary alcohol. And then, there's a bunch of other enzymes that append on some carbohydrate functionality. And then, the whole thing is quickly pumped out of the cell because the cell doesn't want this very dangerous antibiotic hanging around. OK. So, the cell has a complicated machine
37:41
and at the end of it, there's a mechanism that's actually not that well-defined that kind of sweeps up the product and scoots it out the door before it has a time to wreck havoc on the interior of the cell. Let's take a look at the actual polyketide synthase. This is from a structure that was solved
38:00
by my colleague Cheryl Tsai in the chemistry and molecular biology and biochemistry departments here at UC Irvine. And she did this when she was a postdoc and it's really one of my favorite structures of all times and I have a lot of favorite structures but this is a good one. OK. So, in the very center, this is the acyl carrier protein. Oh, OK, big picture first. What we're looking at is we're looking at one
38:22
of these polyketide synthase machines that I showed earlier on a previous slide. So, over here, recall that I showed that there are three of these and I'll show you how they're all linked together in a moment, but each one of these is arranged in the structure of a donut. OK. And in the very center is the acyl carrier protein
38:43
with the phosphopentathione arm that's then going to swivel between each one of these subunits. OK. We now don't think it's entirely a swivel action. It's a little more complicated but I think that's a good model for us to use. So, the first intermediate, the robot arm comes over here
39:00
to the ketone reductase over here. The Claisen condensation reaction is catalyzed and then it goes up to the acyltransferase and then over to the ketone reductase. Oh, sorry, I got this wrong. Ketone synthase at the top, acyltransferase and then ketone reductase and then thioesterase.
39:22
OK. So, these big machines are arranged kind of the way you would arrange the perfect assembly line so that everything is right there and the intermediate can just hop between those different active sites. This is a thing of beauty, isn't it? Check this out. This is the fatty acid synthase that I showed earlier.
39:44
And it turns out that these form dimers. OK. And I'm showing you this because I want to talk to you about how the arrangement of these donut-shaped things into bigger assemblies. OK. So, over here I said that there were three of these donuts. Turns out that these donuts are actually dimers.
40:02
Before I show you that, I need to show you that in fact actually I need to show you that this dimerization is precedent. So, this is the fatty acid synthase that I showed earlier at the very first slide of today's lecture. And what I neglected to tell you was that actually this forms a neat dimer
40:21
where one fatty acid is synthesized on this side and a different fatty acid is synthesized on the other side. So, in similar fashion, oh, and by the way, this structure was solved by Ned and Bond. In similar fashion, it, oh, and it kind of looks like a gingerbread man, right? It kind of looks like a gingerbread man. You see the feet down here, the arms, OK?
40:43
And this is the narrow waist. Anyway, in a similar fashion, the arrangement of these donuts is also in dimer form. OK. So, check this out. OK. So, here's one donut over here. This is module one up here.
41:01
This is module two, module three. And so, what's actually happening is this is actually a dimer of two assembly lines that are wrapped together in double helix fashion. And this one in blue is synthesizing one polyketide and this one in red is synthesizing the same
41:22
polyketide but in parallel. OK. So, in other words, the assembly lines that make these complex chemical structures are actually running all the time and there's multiple assembly lines that are identical that are running right alongside each other. OK. So, this is kind of like going to, I don't know,
41:41
a Mazda factory or something and finding it's not just one assembly line on the floor but two assembly lines that are moving in parallel to each other and both producing the same car at the very end. OK. So, again, this is the, oh, OK. So, again, both polyketide synthases
42:01
and fatty acid synthases are dimers. This one being a circular dimer and this one being a head to tail linear dimer. When things move through the polyketide synthase, the intermediates proceed down this in unidirectional fashion. So, they're starting up here, moving down here, moving down here, moving down here, and they're handed off
42:21
by all those acyltransferases, OK, which then transfers the intermediate to the next acyl carrier protein which then takes it down to the next donut, OK? And then in the end, you end up with this complicated erythromycin over here or erythromycin precursor over here. The fatty acid synthases, as we discussed, however,
42:42
rather than handing things off, instead the same intermediate keeps bopping between each one of those different active sites. And in fact, the intermediate cycles through seven times until you end up with this long chain fatty acid. And at the very end, evidently, it falls off because there's no more room for it to expand, OK?
43:02
It's run out of place in the active site. OK, makes sense? So, you can all draw a diagram to describe how this works and even make some predictions about that sort of thing. You can see how that would be pretty powerful, right? OK. I went to switch gears now, OK? Oh, oh, before I do, any questions about polyketides?
43:24
Anything you want to know about polyketides, that topic? OK. Yeah, Chelsea? Yeah. Yes. Two molecules of this erythromycin precursor.
43:43
Yeah. You know, OK, I can't resist. There's so much more to tell you about this. The one of the really cool areas in this research in this area is to start moving around these active site domains so that in the end,
44:01
you end up programming a different structure. You might imagine setting this thing up so that instead of doing a reduction, maybe you end up with a ketone over here instead. And so, this is a really active and very exciting area of research. So, OK. All right, let's move on. I want to talk to you about a different class of small molecules that are synthesized by cells.
44:24
And these are molecules called terpenes. These are all built from five carbon precursors, OK? And these are familiar things. Geraniol, for example, has that kind of spicy, wonderful flavor of ginger, right? You've all tried that at some point.
44:42
These are things that are really familiar to us. So, polyisoprene is a natural rubber, right? That's actually isolated from these rubber plants. And all of these are built up from five carbon isoprene precursors. Let's just go ahead and count some carbons here, OK?
45:00
So, we're going to have one, two, three, four, five. And then notice that there's a red bond right here. That red bond is going to be joining together these five carbon building blocks. And in the end, these can be strung together in very long polymers, right? So, in the end, you have isoprene, isoprene, isoprene
45:21
and then a very, very long polymer that has that wonderful stretchy feel that natural rubber has, OK? So, let's see. I have to introduce you to some important nomenclature. We're going to call the C10 precursors geranols or geraniols or geraniol.
45:42
And then we're going to call the C15 precursors, that's three isoprenes, farnesyl or farnesol if it's alcohol. And then the C20 precursors, we're going to call geraniol-geraniol alcohol, OK? So, C20 is like two C10s put together, hence the name geraniol-geraniol, OK?
46:01
And I'm going to be using that because it turns out that these are the precursors to much more complex compounds. So, totally analogous to what I was discussing when I was talking about polyketides where we start with these kind of simple linear compounds, terpenes start linear, start simple and then like origami become amazingly
46:22
complex and that complexity is going to be a really great topic for us to discuss. All right, let me show you where we're going with that complexity. So, up at the top, these are the straight chain precursors that I showed on the previous slide, geraniol pyrophosphate, farnesyl pyrophosphate and geraniol-geraniol pyrophosphate.
46:42
These again are synthesized from five carbon building blocks, two building blocks, isopentenyl pyrophosphate and dimethyl allyl pyrophosphate. I'll show you those two in closer detail in a moment. So, let's suspend for a moment where they come from. These guys over here,
47:00
these straight chain precursors can be folded up by a class of enzymes known as terpene synthases or terpene cyclases and the net effect is you get out these very complex skeletons of really complicated molecules. Okay, so for example, this is the skeleton that eventually will become taxol down here.
47:22
Taxol results from a series of tailoring enzymes that oxidize various carbons in this taxodyne framework and then append on things like this little short peptide fragment, this dipeptide fragment. Okay, so you start off simple, you start off linear
47:41
and then you get increasingly, you fold things up, you cyclize and then you modify after it's cyclized. Okay, and what's great about this is this follows the same formula that I gave you when we talked about the fatty acids. You start off simple, you start two carbon, three carbon building blocks, you build linear things and then you cyclize them and then you get more complicated.
48:03
Okay, exact same formula here except now we're going to build everything out of hydrocarbons. There's going to be very few oxygens. Oh, and by the way, these are familiar compounds as well. Limonene, this is isolated. I think you guys do this isolation, right, from like orange peel.
48:21
You did this back in Chem 51A lab or something like that or 51B lab. Yeah, so this is the compound that you isolated. You'll learn how this is synthesized by cells. It can also be oxidized to give menthol, a nice wintergreen kind of taste, a nice minty taste.
48:41
This amphoradiene is modified to make artemisinin. This is a compound that has important anti-malarial properties and is extremely useful therapeutically. All right, this is kind of the big picture. Everyone with me so far? All right, let's zoom down and start looking at the nuts and bolts.
49:01
Oh, more big picture, sorry. The enzymes that make the, that do the cyclization have remarkable, remarkable chemical specificities, okay. So, starting from the same precursor, different terpene synthases can direct the synthesis of different stereoisomers of this bicyclic compound
49:25
and then other terpene synthases can direct whether or not you get spirofused rings. This is a spirofused ring. When two rings are joined at a quaternary carbon that's called spirofused. These are called, you know, cis or transfused
49:40
and you can even get monster rings. So, whether or not you get a 5-7 or a 6-6 ring or larger rings is all controlled by these terpene synthases and they're really, really good at what they do. Okay, so this is, they are controlling stereochemistry and regiochemistry in water at room temperature.
50:03
Okay, I've already talked about other, there's lots of other complex terpenes I can show you. This is an absolutely fascinating class of molecules. There's targets for synthesis, et cetera. Let's talk about how they're put together starting at the most basic level down here. So, I told you that all of these are synthesized
50:21
from five carbon building blocks that are going to be isoprenes in the finished structure, okay, where an isoprene is this compound here of the five carbons with the carbon-carbon double bond in the middle. Okay, what happens is there's one starting material is this
50:42
dimethyl allyl pyrophosphate. This has an allylic pyrophosphate as its leaving group. And no surprise, the active site that's going to catalyze this reaction has, of course, a magnesium available to act as a Lewis acid and stabilize the pyrophosphate, making it a good leaving group.
51:03
Okay, and the product here is going to be the ultra-stable allylic carbocation, okay? So, what happens is the allylic carbocation forms. This can form resonance structures, hence its stability. The resonance structure will be if the cation, sorry, the tertiary carbocation, but it's set up in a way,
51:22
the tertiary carbocation is held away from the incoming nucleophilic isopentenyl pyrophosphate, this compound here, and this sets you up for a nucleophilic attack, which then gives you a new tertiary carbocation.
51:41
And then in the final step, a beta elimination step gives you in the end this geranil diphosphate, okay? And then you can imagine this could be the starting material for additional IPPs to be added on using exactly the same chemistry that I'm showing you here.
52:00
Okay, make sense? All right, let's take a closer look. All right, so it turns out that these things are synthesized using reactions that are, you know, completely understandable. So now, I'm stepping back and I'm showing you how to synthesize these compounds down here.
52:22
IPP and dimethyl allyl pyrophosphate. First, these two can result from a simple isomerization, right, that's this carbon-carbon double bond over here getting isomerized to form the more stable, the more substituted olefin of dimethyl allyl pyrophosphate,
52:41
okay, and there's an enzyme that catalyzes that reaction, okay, and active site looks like this. It has a manganese ion that can form a Lewis acid relationship with the glutamate in the active site and this can act
53:01
as a base to catalyze this deprotonation over here and then there's a nearby bile that acts as an acid to protonate the resulting carbanion as this thing gets deprotonated.
53:20
Okay, let's start at the beginning, okay, so everyone, this reaction down here make sense? Okay, so let's talk about how, so if we can get to here, then we can get to here. How do we get to this IPP? Okay, IPP is synthesized using more or less the same sort of chemistry that I've shown you previously in the context
53:43
of fatty acid synthesis, okay, it's basically going to be set up using a series of Claisen and Aldol condensations, okay, so all carbon-carbon forming events are going to use the Aldol reaction and there's an enzyme called HMG-CoA synthetase that's going
54:02
to synthesize this HMG-CoA molecule over here, okay, from HMG-CoA you can then readily get to this compound here, okay, using phosphorylation from ATP and then, so you hydrolyze this ester, reduce the,
54:22
or sorry, hydrolyze this ester and then reduce it down to the alcohol and then ATP then transfers pyrophosphate to the primary alcohol and then you can do an elimination reaction, okay, so this is a decarboxylation and then elimination of alcohol,
54:40
does anyone remember the context where we saw a reaction like this on Tuesday? We've seen the mechanism of this reaction before, do you recall when we saw it? On Tuesday, decarboxylation giving us an enol or enolate,
55:10
do you remember that, what was the sequence, why did we do that?
55:29
Yeah, so we did this decarboxylation, right, and that gave us an enolate and then what was the enolate used for, do you remember? It was the Claisen, right, so this is being done
55:42
by ketosynthase, right, so this is totally analogous chemistry, nothing new here, okay, thanks Carl, you rescued us. Okay, then the thing is this reaction over here when mevalonic acid is synthesized, oh, here's the reduction here by HMG-CoA reductase,
56:03
this mevalonic acid then becomes a precursor for the eventual synthesis of cholesterol, so if you could shut down this step or this step, you'd have a way of preventing formation of cholesterol and that actually is pharmaceutically a very important reaction, this is a reaction
56:21
that is very extensively inhibited, so HMG-CoA reductase inhibitors include compounds like Lipitor, Mevacor, et cetera, and many of these start to look a little bit like the mevalonic acid, right, so this compound here is kind of like a folded up mevalonic acid
56:40
and then up here you can imagine hydrolysis of this lactone will give you something that looks a little bit like mevalonic acid as well, so these compounds are very widely prescribed, it's literally billions upon billions of dollars a year of this stuff is sold, probably everyone you know over the age of 50 is on one of these compounds because they're so effective at suppressing formation
57:02
of cholesterol and that seems to have such positive benefits for cardiovascular health and also even some anti-cancer abilities as well, so you know, these are remarkable compounds, they've changed how we treat patients and they're sold in enormous volumes.
57:21
Okay, and again these things look like the product, so they work as product mimics. All right, so I've already shown you how to connect up the isoprene units and one thing I need to point out is that this is an example of a very rare non-Aldol carbon-carbon bond
57:41
forming reaction, this is not very typical and that makes it kind of special I think. Okay, all right, last mechanism of the day, this one blows me away, there's a class of it, the next step here is to take this straight chain precursor
58:02
and then fold it up into some semblance of the rings and then cyclize things, so the class of enzymes that does this again are called terpene synthases and here's one example of this, a protein called the ristalochine synthase, this starts with varnasil pyrophosphate,
58:21
the way this works is that the enzyme has a very deep active site, it's kind of like a deep cave, okay, so deep, deep active site, that's shown here, right here at the mouth of the cave, this is actually a transition state analog, a product analog,
58:43
or sorry, substrate analog that the enzyme cannot act upon, so when the crystallization took place this shows us where the active site is, okay, and again the active site is this really deep cave, now at the tips of my fingers over here there's a series of carboxylate bearing side chains
59:01
that can then chelate magnesium ions, okay, those magnesium ions as you might expect form a Lewis acid relationship with the pyrophosphate, analogous to what we saw a couple of slides ago, okay, similar chemistry, the fact though is that this draws the mouth of the cave shut, the magnesiums come
59:24
down and the pyrophosphate gets coordinated to those magnesiums and that draws the cave shut, now the interior of the cave, the kind of the back side in here is highly hydrophobic, this makes sense,
59:40
right, the molecule that has to bind is very hydrophobic, it has 15 carbons in a row, so this side over here that's going to be at the mouth of the cave binding to the magnesium ions is really nicely hydrophilic and then this side over here is hydrophobic, so what happens is as
01:00:00
the cave shuts, all of the water in the cave gets squeezed out, OK? So all of the water gets pushed out. This will be critical in a moment and I'll tell you why that is in a second, OK? Now, OK, so we've seen the binding by the farnesyl pyrophosphate. The second step here is in the active site.
01:00:21
The active site is shaped in such a way that the C15 piece is pushed into some semblance of the correct product. So the one enzyme over here that's going to be synthesizing a risk of low chain will have an active site that's shaped to push this straight chain precursor
01:00:43
into these two rings. But the one that's going to be synthesizing say this spirofuse ring is going to start the same starting material but in this case, it's going to be pushing the farnesyl into something that looks like this, OK? Next step here is, again, to clamp down the diphosphate
01:01:01
with the two magnesium ions and push out the water. The magnesium ions, again, act as Lewis acids and they trigger the dephosphorylation reaction. They stabilize the pyrophosphate leaving group allowing it to take off and that has the effect of triggering a carbocation cyclization
01:01:21
that eventually leads you down to one and only one product like this one. Now, the complicated thing here is that this carbocation cyclization is not some simple one stepper. This is a really complicated reaction. I'll show you what it looks like on an upcoming slide but it involves multiple steps and it involves a requirement
01:01:43
to stabilize carbocations preferentially, carbocation intermediates preferentially. And so the enzyme has evolved this active site that has, for example, aromatic residues positioned neatly over carbocations to stabilize those carbocations by cation pi interactions, the same interactions
01:02:03
that we saw very early in this course when we talked about non-covalent binding. In addition, many of these compounds are synthesized through other intermediates and the active site has ways of actually then catalyzing reactions on those intermediate compounds as well.
01:02:23
Oh, and then the final step, of course, is the beta elimination to produce the product and this is analogous to the beta elimination that we saw earlier to where the farnesyl pyrophosphate was synthesized through an elimination reaction. This is that same step. OK. So these are truly wondrous reactions and these are things
01:02:44
that we can talk about quite a bit. I would love to talk to you about them some more but I actually want to reward all of you who have shown up today with a very, very short and easy quiz. So I'm going to stop here actually. So, please take out a half sheet of paper and get ready for a quiz.