BIOL 302 1st Edition Lecture 26Outline of Last Lecture I. GlycolysisOutline of Current Lecture I. Terminal electron acceptorII. Citric acid cycleIII. Lipotic acid IV. Citrate synthaseV. Replenishing oxaloacetateVI. Aconitase isomerizesCurrent I. Moving toward O2 as a terminal electron acceptorA. Glycolysis partially oxidizes glucose – much energy is left to harvestB. Fermentation does not use oxygen as a terminal electron acceptorC. Most of the ATP generated from glucose is provided by aerobic processesD. Further oxidation strips more electrons from pyruvate to power oxidative phosphorylation; the citric acid cycle sets this upII. The Citric Acid Cycle Takes Place in the MitochondrionA. Glycolysis occurs in the cytoplasm, but the enzymes for the citric acid cycle are found in the mitochondrial matrix (except for succinate dehydrogenase, which is in the inner mitochondrial membrane)B. The two-membrane structure allows for the set-up of a proton gradient, with high concentration in the intermembrane space.These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.III. The Citric Acid CycleA. harvesting of high-energy electrons from carbon fuelsB. Oxidizes an acetyl group to two molecules of carbon dioxide -CO-CH32 CO2C. Does not generate much ATP, nor does it involve O2IV. CA cycle generates electron carriers for cellular respirationA. Goal of the citric acid cycle is to transfer electrons from carbon fuels to reduced electroncarriers for the eventual synthesis of ATPB. Oxidative phosphorylation makes ATP via the reduction of O2 to generate a proton gradientV. The cycle has three different namesA. Citric Acid Cycle1. Named for citrate, the first product produced after a new 2-carbon unit is introducedinto the cycleB. Krebs Cycle1. Named after Hans Adolf Krebs for seminal work with Albert Szent-Gyorgyi in the 1930sC. Tri-Carboxylic Acid (TCA) Cycle1. Citrate is a tricarboxylic acidVI. Pyruvate from glycolysis enters the cycle as Acetyl-CoAA. Notice that CO2 and electrons are released at this step.VII. Pyruvate Dehydrogenase ComplexVIII. Pyruvate Dehydrogenase ComplexA. Pyruvate + CoA + NAD+ Acetyl-CoA + CO2 + NADH + H+B. Also requires the catalytic cofactors: thiamine pyrophosphate1. (TPP, step 1),2. lipoic acid (step 2)3. flavin adenine dinucleotide (FAD, step 3)IX. The PDH complexA. Multiple Cofactors are involved1. Catalytic2. StoichiometricCoenzyme ANAD+X. Step 1. DecarboxylationA. pyruvate dehydrogenase– E11. The structure of TPP makes the ring proton much more acidic than normal2. The carbanion attacks the pyruvate carbonyl group, creating C-C bond3. This promotes decarboxylation, breaking a C-C bond4. The product then rearranges to form hydroxyethyl-TPPXI. Step 2. OxidationA. pyruvate dehydrogenase– E11. The hydroxyethyl group is oxidized to acetate and transferred to lipoamide2. Also catalyzed by the PDH E1 subunit. Forms an energy-rich thioester bond.XII. Step 3. Formation of Acetyl-CoA and RegenerationA. Transfer - Dihydrolipoyltransacetylase – E2B. Regeneration - Dihydrolipoyl dehydrogenase – E3C. The regeneration step yields NADH for oxidative phosphorylationXIII. The PDH complexA. derivative of lipoic acidB. -linked to a lysine residue by an amide linkageXIV. Lipoic acidA. Organosulfur compound derived from octanoic acid (C8)XV. The many chemistries of PDH are allowed by the flexibility of the lipoamide prosthetic groupXVI. A series of reactions catalyzed by the flexible lipoamide groupA. TPP intermediate generated deep within E1B. E2 inserts the lipoamide arm into E1C. E1 catalyzes transfer of an acetyl group to the lipoamide which enters E2 (pink) active siteD. Acetyl moiety transferred to CoA, producing acetyl CoA 5. Now at the E3 active site, lipoamide is oxidized by FADE. Reactivated lipoamide is ready to begin another cycleF. Final reaction product, NADH, is produced. FAD is regeneratedXVII. Clicker Question: Which of the following best describes the biochemical role of coenzyme A?A. It activates acetyl groups for group transfer.B. It assists in the transport of metabolic intermediates across membranes.C. It shuttles electrons within the electron transport chain.D. It introduces adenine nucleotides into metabolic products.E. It introduces thiol groups into metabolic products.XVIII. Clicker Question: In cells, NADH serves as a carrier of which of the following?A. Phosphoryl groups B. Acetyl groupsC. Hydroxyl groupsD. Methyl groupsE. ElectronsXIX. Oxidation of 2-carbon unitsXX. Citrate SynthaseA. Gateway to the citric acid cycleB. Condenses 2-carbon acetyl-CoA with 4-carbon oxaloacetateC. This reaction is an aldol condensation followed by hydrolysis of the CoA thioesterXXI. Citrate SynthaseA. As we’ve seen in other mechanisms, an enol intermediate is created through the sequential action of specific bases and acids in the enzyme active siteB. Citrate synthase undergoes major conformational rearrangements throughout catalysis to maintain a strictly sequential reaction that prevents hydrolysis of the thioester until the condensation is completed, then releases CoA-SHXXII. Aconitase isomerizes citrate in 2 stepsA. Citrate is then isomerized to form isocitrate – a hydroxyl group is moved to the adjacent carbonB. Dehydration followed by re-hydration across the resulting double bondC. Positions the hydroxyl group to be oxidized and participate in decarboxylation in the nextstepXXIII. Aconitase is a non-heme iron-sulfur proteinA. Four iron atoms are complexed to four inorganic sulfides and three cysteine sulfur atoms, leaving one iron atom available to bind citrate and then isocitrate through their carboxylate and hydroxyl groupsXXIV. Isocitrate Dehydrogenase: oxidative decarboxylationA. Isocitrate is oxidized to yield one molecule of NADHB. We see decarboxylation to drive the formation of an enolate intermediateXXV. -ketoglutarate dehydrogenaseA. Second round of oxidative decarboxylation in the citric acid cycle (redox)B. Reaction mechanism is entirely analogous to pyruvate dehydrogenase and also involves three enzyme activitiesC. Yield is one more reduced electron carrier (NADH). Also requires TPPXXVI. Succinyl-CoA SynthetaseA. Named for the reverse reaction, succinate → succinyl-CoAB. The only step in the citric acid cycle that directly yields a high phosphoryl-transfer potential molecule (ATP or GTP). C. Accomplishes this by