biochem: Lecture 2 TCA Cycle (1-30)

does acetyl-coa formation from pyruvate happen in cytosol or mitochondria
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describe step 2 of pyruvate dehydrogenation and decarboxylationthe hydroxyethyl group is oxidized to the level of carboxylic acid (acetate). The two electrons removed reduce the s-s of lipoate on E2 to two thiol groups and then one of the thiols are esterified to have an acetyl group. Then the acetyl group is transferred from the thiol on lipoate to CoA to form acetyl-CoAdescribe step 3 of pyruvate dehydrogenation and decarboxylationThe rest of the transfers are electron transfers and they occur to regenerate the oxidized disulfide form of the lipoyl group of E2 to prep the complex for another round of oxidation the electrons the lipoyl group gained from the transfer of acetyl to Coa are passed to FAD that is in E3 and FAD becomes FADH2. FADH2 passes its electrons to NAD+ to make 1NADH and Lypoyl group is oxidized and ready to pick up another acetylgive two examples of electron channeling1. e channeling between glyceraldehyde-3-P dehydrogenase and phosphoglycerate kinase 2. pyruvate dehydrogenase between E1,E2, and E3what is the input and output of the TCA cycleinput: 1 acetyl-Coa output: 3NADH, 1FadH2, 1GTP, 2CO2what is the first regulatory step of the TCA cycleacetyl-coa + oxaloacetate -> citratewhy is the first step of the TCA cycle exergonichydrolysis of acetyl-Coafour of the eight steps of the TCA are oxidations. What are the four output products of each of these oxidations3NADH and 1FADH2what molecule of the TCA cycle acts catalytically?oxaloacetate because it is consumed and reproduced so theoretically it is not consumed like an enzymeone molecule of oxaloacetate can theoretically bring about oxidation of how many acetyl groups?infinite amount of acetyl groupslist the names of all 8 steps of the TCA cycle1. aldol condensation 2.dehydration/rehydration 3. oxidative decarboxylation 4. oxidative decarboylation 5.substrate level phosphorylation 6. dehydrogenation 7. hydration 8. dehydrationtrue or false, the carbons released from the the TCA cycle came originally from acetyl-CoafalseTCA cycle step 1 1. reactant product 2. enzyme 3. reversibility + free energy 5. chemical logic1. acetyl coa + oxaloacetae -> citriate 2. citrate synthase 3. non-reversible very exergonic 4. CoA-SH 5. citrate made in prep for oxidative decarboxylationwhy is step 1 of TCA exergonichydrolysis of acetyl-Coawhy does binding order matter for citrate synthasefirst oxaloacetate binds and induces an open form of the enzyme that can now accept acetyl-Coa acetyl-CoA now binds and causes the enzyme to close for optimal catalysis (more compact)know mechanism of citrate synthasepractice on paperTCA cycle step 2 (a and b) 1. reactant product 2. enzyme 3. reversibility + free energy chemical logic?a) 1. citrate -> cis-aconitate 2. aconitase 3. reversible endergonic 4. water b) 1. cic-aconitate -> isocitrate 2. aconitase 3. reversible endergonic 4. none chemical logic: to move the hydroxyl to C-4 to allow for a carbonyl to form on C-4 inn the next step to make formation with Mn2+what mediates the catalysis of citrate to isocitrate in the enzyme aconitasebasic residue and iron-sulfur centreif citrate to isocitrate is unfavorable, how does it move forwarddriven forward because isocitrate is rapidly consumed in next stepTCA cycle step 3 1. is isocitrate oxidized or reduced into alphaketoglutarate? 2. enzyme 3. metal catalysis 4. why is Mn so important? 5. reversible and energy1. oxidized 2. isocitrate dehydrogenase 3. Mn2+ in active site of enzyme 4. Mn creates a highly charged electrohile that takes the electron from the carboxyl group allowing it to leave as a CO2 5. not reversible and very exergonicwhat is the first oxidative decarboxylation step of TCAisocitrate -> alphaketoglutaratewhat is the intermediate of isocitrate -> alpha-ketoglutarateoxalosuccinatewhat is the second regulated step of tcaisocitrate to alpha ketoglutaratedescribe the chemical logic of isocitrate to alpha ketoglutarate1. isocitrate oxidized by NAD+ or NADP+ to make oxalosuccinate intermmediate 2. decarboxylation occurs that is facilitated by bound Mn2+ 3. rearrangement of the nol generates alpha ketoglutaratewhy does isocitrate dehydrogenase catalyze the oxidation before the decarboxylationOxidation creates a carbonyl group which can then stabilize thenegative charge resulting from decarboxylationTCA cycle step 4 1. enzyme 2. product 3. co-substrates 4. energy and reversibility1. alpha ketoglutarate dehydrogenase complex 2. succinyl-CoA 3. CoA-SH and NAD+ 4. very exergonic and nonreversiblewhat TCA cycle reaction is identical to pyruvate dehydrogenase rxn?alpha ketoglutarate dehydrogenase complex to make succinyl-CoAwhat is the name of the rxn to make succinyl-CoAoxidative decarboxylation processwhat is the third regulatory step of TCAproduction of succinyl-CoAdraw out the mechanism of the creation of succinyl-CoA like pyruvate dehydrogenase reactiondraw it outin what ways are alpha-ketoglutarate dehydrogenase and pyruvate dehydrogenase similarboth have similar E1, E2, and E3 domains and both have TPP, lipoic acidTCA cycle step 5 1. product 2. input and output co-substrates 3. reversibility and energy 4. enzyme1. succinate 2. GDP + Pi input and GTP and CoA-SH output 3. reversible and exergonic but basically zero 4. succinyl-CoA synthetasewhy is the net energy of the production of succinate basically zero?because the free energy released from the hydrolysis of the thioester is used to make GTPwhat is the name of the reaction of making succinatesubstrate level phosphorylationdescribe the steps of the production of succinate1. succinyl-CoA is phosphorylated by the hydrolysis of thioester bond and Coa-SH is released 2. succinyl phosphate is now enzyme bound and the phosphate is trasnfered to a Histidine group in the active site which allows succinate to leave 3. the phosphate bound to the histidine now can phosphorylate GDP and succinyl-CA synthetase active site is ready for another succinyl-CoA to enterTCA cycle step 6 1. product 2. enzyme 3. reversibility and energy 4. co-substrate1. fumarate 2. succinate dehydrogenase 3. reversible and energy zero 4. FADwhen fumarate is made, was succinate oxidized or reduced?oxidizedwhich TCA cycle enzyme is not able to diffuse in the mitochondrial matrixsuccinate dehydrogenasewhat is the first reaction in the TCA cycle that makes an electron carrierformation of fumarate because FADH2 is madeTCA cycle rxn 7 1. intermmediate 2. product 3. reversibility and energy 4. input 5. enzyme1. fumarate carbanion 2. l-malate 3. reversible and exergonic but zero 4. hydroxyl and proton 5. fumarasewhat type of double bond does fumarate havetransfumarate and maleate both have double bonds and two carboxyl groups. Fumarate is trans double bond and maleate has a cis double bond. Can fumarase add water across the double bond of both molecules?no. Fumarase adds water to the c=c only to the trans double bondif fumarase could process maleate (that has a cis-double bond instead of trans like in fumarate) what would be the substrate product?D-malateTCA cycle rxn 8 1. product and side product 2. enzyme 3. name of chemical reaction 4. reversibility and energy1. oxaloacetate and NADH 2. malate dehydrogenase 3. ox 4. reversible but VERY endergonicdescribe why the production of oxaloacetate from l-malate can occur despite it being very unfavorableoxaloacetate is continuously consumed by the starting rxn of TCA when citrate is made. In this step, acetyl-CoA is hydrolyzed and this is extremely exergonic which contributes to funneling oxaloacetate into the reaction additionally, oxaloacete is kept at very low concentrations due to its constant removaluse the equation deltaG=deltaG + RTln(OA/M)describe the general chemical logic from alphaketo to oxaloacetatealphaketo is just like succinyl-Coa only succinyl-Coa has CoA instead of COO- like in alphaketo sucinyl-CoA is made into succinate by removal of CoA and this sets it up to become oxaloacetate succinate is just like fumarate only two H are removed on succinate 1 on C2 and 1 on C3 to get a double bond between the two carbons. This allows fumarate to have a trans double bond the trans double bond of fumarate allows fumarase to make it inot L-Malate which is just like succinate only it now has a hydroxyl group on C-3 the hydroxyl on C of l-malate allows for a carbonyl to be made on C to make oxaloacetatewhat four steps are oxidations in TCAalpha-keto production succinyl-CoA production fumarate production oxaloacetate productionhow many ATP are made from the TCA cycle (dont count two pyruvate)3NADH=7.5ATP FADH2=1.5ATP GTP=1ATP total: 10from glycolysis to TCA calculate the ATP made as an activity (slide 30)which TCA step is an aldol condensationproduction of citratewhich TCA step(s) are dehydration/hydrationproduction of isocitrate production of malatewhich TCA steps are dehydrogenationproduction of fumarate production of oxaloacetatewhich TCA cycle steps are oxidative decarboxylationproduction of alphaketo production of succinyl-Coawhat step of TCA is substrate level phosphorylationproduction of succinateacetyl-Coa must be completely oxidized to CO2 to maximize the energy to be extracted. Why cant we directly oxidize acetate to CO2?decarboxylation of the two carbon aciid would yield CO2 and methane if we direclty oxidized acetyl-CoA to CO2. methane is harmful to produce in the human body due to lack of cofactors that can convert it.why are carbonyl groups important for metabolic pathways?the carbon in a carbonyl group is partially positive and therefore an electrophilic centre which can make a carbanionwhat solves the problem of methane being produced by acetyl-Coa in the TCA cyclewhen acetylcoa reacts with oxaloacetate, the oxaloacetate carbonyl acts as an electrophilic center which is attacked by the methyl carbon of acetylcoagive an example of an electrophilic attack in the TCA cyclein step 1, the methyl carbon oc acetyl-CoA is an electrophile that attacks the electrophilic center of the oxaloacetate carbonyl carbonoxaloacetate is very low concentration. What about rxn 1 of the TCA helps the rxn happenthe hydrolysis of the thioester makes the forward reaction of rxn 1 very exergonic and this allows the for the reaction to occur despite the low concentration of oxaloacetatewhat intermediate in the tca cycle is a tricarboxylic acid? what is also a tricarboxylic acidcis-aconitate isocitrate citrateisocitrate is at a low concentration in the TCA cycle, what pulls it forward?that it is immediately consumed in the next stepmalonate is a strong competitive inhibitor of succinate dehydrogenase. what does its activity hault in the cell? 1. TCA 2. electron transport chain 3. ATP synthesis 4. glycolysis 5. TCA and ETC1. it will only hault the TCA cyclewhy is oxaloacetate kept at a low concentrationbecause its formation from L-malate, the equilibrium lies far to the left which pulls the reaction toward the formation of oxaloacetate to keep the equilibrium balanced. So as oxaloacetate is consumed, more is made to maintain the far left equilibrium