-
· Conversion of pyruvate into acetyl CoA by __ is the link between __ and __ because __ is the fuel for the CAC; all fuels are metabolized to __ or components of the CAC
· The CAC begins with the __--> __ and __are the products
- pyruvate dehydrogenase complex
- glycolysis and cellular respiration
- acetyl CoA
- acetyl CoA
condensation of a four-carbon unit, oxaloacetate, and a two-carbon unit, the acetyl group of acetyl CoA
citrate and CoA
-
o Citrate synthase catalyzes this __ followed by __
§ Oxaloacetate first condenses with acetyl coA to form __, a molecule that is energy rich because it contains the __ that originated in acetyl CoA
· Hydrolysis of __to __ and __ drives the overall reaction far in the direction of the synthesis of citrate
o The hydrolysis of the __powers the synthesis of a new molecule from two precursors
- aldol condensation followed by hydrolysis
- citryl CoA
- thioester bond
- citryl CoA thioester
- citrate and CoA
- thioester
-
· Citrate synthase prevents __
o Each active site of citrate synthse (a dimer) is located in a __between the large and small domains of a subunit, adjacent to the subunit interface
o The enzyme undergoes __ in the course of catalysis
§ Citrate synthase exhibits __; __binds first, followed by __
wasteful hydrolysis of acetyl CoA
cleft
large conformational changes
sequential, ordered kinetics
oxaloacetate
acetyl CoA
-
· Oxaloacetate induces a major structural rearrangement leading to the __
o The binding of oxaloacetate does what?
§ In each subunit, the small domains do what?
• Movements as large as __ are produced by the __ elicited by quite small shifts of the __
o These structural changes create a __
- creation of a binding site for acetyl CoA
- converts the open form of the enzyme into a closed form
- rotates 19 degrees relaive to the large domain
- 15 angstroms
- rotation of alpha helices
- side chains around bound oxaloacetate
binding site for acetyl CoA
-
· Citrate synthase catalyzes the __ by doing what?
o The __ and __ of __ transforms acetyl CoA into an __, which goes on to __.
§ The newly formed __ induces more structural changes in the enzyme,c ausing the active site to become __. The enzyme cleaves the __ by __. __leaves the enzyme, followed by __, and the enzyme returns to the initial open conformation.
- condensation reaction
- bringing the substrates into close proximity, orienting them, and polarizing certain bonds
- donation and removal of protons
- enol intermediate
- attack oxaloacetate
- citryl CoA
- completely enclosed
- citryl CoA thioester
- hydrolysis
- CoA
- citrte
-
· Essentially, citrate synthase can hydrolyze __ but not __. It discriminates against the two because, first, __; and, second, __. This is __ again
- citryl CoA
- acetyl CoA
- acetyl CoA does not bind to the enzyme until oxaloacetate is bound and ready for condensation
- the catalytic residues crucial for the hydrolysis of the thioester linkage are not appropriately positioned until citryl CoA is formed
- induced fit
-
· Because the hydroxyl group is not in a right place, citrate is __into __to enable the six-carbon unit to undergo __
o The __ is accomplished by a __ followed by a __. The result is the interchange of an H and an OH
§ The enzyme catalyzing both steps is called __, which is an __ protein that contains iron not bound to heme
- isomerized
- isocitrate
- oxidative decarboxylation
- isomerization of citrate
- dehydration step
- hydration step
- aconitase
- iron-sulfur
-
· Its four iron atoms are bound to __ and __, leaving one __ available to bind citrate through one of its COO- groups and an OH group.
o This __ participates in __ and __ the bound substrate
- four inorganic sulfides
- three cysteine sulfur atom
- iron atom
- Fe-S cluster
- dehydrating and rehydrating
-
· The first of four oxidation-reduction reactions in the CAC is catalyzed by __; it is the __ of __. Int his reaction, __is lost and __ is formed.
o This first oxidation generates the first __
- isocitrate dehydrogenase
- oxidative decarboxylation
- isocitrate
- CO2
- alpha-ketoglutarate
- high-transfer- potential electron carrier, NADH
-
· The second __ is the second step, the formation of __ from __, which is catalyzed by __, an organized assembly of three kinds of enzyme that has a similar mechanism to the pyruvate dehydrogenase complex
o Both reactions include the __of an __ and the subsequent formation of a __ with __that has a __.
- oxidative decarboxylation
- succinyl CoA
- alpha-ketoglutarate
- alpha-ketoglutarate dehydrogenase complex
- decarboxylation
- alpha-ketoacid
- thioester linkage
- CoA
- high transfer potential
-
· __ is an energy rich thioester compound. In the __ reaction, the cleavage of the __ powers the synthesis of the __ from the __ and the __
o The cleavage of the __ is coupled to the __
§ This reaction is readily reversible and is catalyzed by __
- Succinyl CoA
- citrate synthase
- thioester bond
- six-carbon citrate
- four-carbon oxaloacetate and the two-carbon fragment
- thioester bond of succinyl CoA
- phosphorylation of a purine nucleoside diphosphate, usually ADP
- succinyl CoA synthetase
-
· This reaction is the only step in the CAC that yields a compound with __
o In mammals, there are two __ of the enzyme, one for __ and one for __
§ __ have the GTP one; __have the ATP form
· The GDP requiring enzyme works in the __; used to power __
- high phosphoryl-transfer potential
- isozymic forms
- ADP and one for GTP
- Anabolic tissues
- tissues that respire a lot
- reverse of the TCA cycle
- synthesis of succinyl CoA
-
· The enzyme __ catalyzes the reaction: GTP+ ADPßà GDP + ATP
· The mechanism of __ has energy in the __; and, this energy is transformed into __
o Step one: __ by __, which generates __
§ A histidine acts as a __that does what?
- nucleoside diphosphokinase
- succinyl Coenzyme A synthase
- thioester molecule
- phosphoryl-group transfer potential
- displacement of CoA
- orthophosphate
- enery rich succinyl phosphate
- moving arm
- detaches the phosphoryl group, swings over to the bound nucleoside diphosphate, and transfers the group to form the NTP
-
· __ is an __; a phosphoryl group is transferred first to __ bound in the __ and then to an __ in the __
o Each subunit has two domains
§ The amino-terminal domains have different structures
· Alpha forms a __, binding __ of succinyl CoA synthetase
· Beta is an __, which binds and activates ADP
- Sucinyl CoA synthetase
- a2B2 heterodimer
- succinyl CoA
- alpha subunit
- NDP bound
- beta subunit
- Rossmann
- ADp
- ATP-grasp domain
-
· Reactions of four-carbon compounds is the final stage: __
o A __ is converted into a carbonyl group in three steps: __
§ __is regenerated and more enery is extracted as __ and __
- reneration of oxaloacetate
- methylene group
- oxidation, hydration, and second oxidation
- Oxaloacetate
- FADH2 and NADH
-
· Succinate is oxidized to __by __, passing electrons to __because the free-energy change is insufficient to reduce NAD+.
o FAD is always the electron acceptor in oxidations that remove __ from a substrate
§ In __, the isoalloxazine ring of FAD is covalently attached to a histidine side chain of the enzyme
- fumarate
- succinate dehydrogenase
- FAD
- two hydrogen atoms
- succinate dehydrogenase
-
o Succinate dehydrogenase is an __ proteins ahd have __
§ It is in the __ and directly associated with the __, the link between the CAC and ATP formation
· __produced by oxidation of succinate like NADH in other reactions; instead, __ are transferred from __directly to __of the enzyme, which passes the electrons to __, which passes them to xygen
- iron-sulfur
- 3 different iron-suulfur clusters
- inner mitochondrial membrane
- eTC
- FADH2
- two electrons
- FADH2
- iron-sulfur clusters
- CoQ
-
· The next step is the __.
o __catalyzes the __ addition of H+ and OH-. The OH- group adds to only one side of the double bond of fumarate. So, only the __ is formed
· Finally, __is oxidized to __by __and __ is the hydrogen acceptor
- hydration of fumarate to form L-malate
- Fumarase
- stereospecific trans
- L isomer
- malate
- oxaloacetate
- malate dehydrogenase
- NAD+
-
· The net reaction of the CAC is:
o Acetyl CoA + 3 NAD+ + FAD + ADP + Pi + 2 H2O à 2 CO2 + 3 NADH + FADH2 + ATP + 2 H+ + CoA
-
Summary
o Two carbon atoms enter the cycle and condense with __. Two carbon atoms leave the cycle in the form of __int eh successive __ catalyzed by __ and __
o __ leave the cycle in four oxidation reactions. Two __molecules are reduced in the oxidative decarboxylations of __, one __molecule is reduced in the __, and one __ is reduced in the oxidation of __
- oxaloacetate
- CO2
- decarboxylations
- isocitrate dehydrogenase
- alpha-ketoglutarate dehydrogenase
- Four pairs of hydrogen atoms
- NAD+
- isocitrate and alpha-ketoglutarate
- FAD
- oxidation of succinate
- NAD+
- malate
-
o One compound with __, usually ATP, is generated from the cleavage of the __
o Two __ are consumed: one in the __ and the other in the __
- high phosphoryl-transfer potential
- thioester linkage in succinyl CoA
- water molecules
- synthesis of citrate by the hydrolysis of citryl CoA
- hydration of fumarate
-
· __ revealed that the two carbon atoms that enter each cycle are not what, but what?; they remain __
o The two carbons that enter the cycle will be released as __in subsequent trips through the cycle
·
- Isotope-labeling studies
- not the ones that leave, but instead are retained during the initial two decarboxylation reactions
- incorporated in the four-carbon acids of the cycle
- CO2
-
Evidence shows that the enzymes of the CAC are __, enhancing the efficiency of the CAC due to __; the structure is called a __
physically associated with one another
substrate channeling
metabolon
-
· The ETC oxidizes the __ and __ formed in the CAC. The transfer of electrons from the carriers to oxygen leads to the __ across the __; the __ powers the generation of ATP
o 2.5 ATP per NADH; 1.5 ATP per FADH2
- NADH and FADH2
- proton gradient
- inner mitochondrial membrane
- proton-motive force
-
§ __ are generated when the ETC oxidizes __ and __, and one high-transfer potential phosphoryl group is directly formed in one round of the CAC
· Thus, one __ generates approximately 10 molecules of ATP
Nine high-transfer potential phosphoryl groups
three NADH molecules and 1 FADH2 molecule
acetyl unit
-
· The CAC cycles only under aerobic conditions because __ can be regenerated in the mitochondrion only by the __
o Glycolysis has both an aerobic and anaerobic mode, whereas the CAC is strictly __
- NAD+ and FAD
- transfer of electrons to molecula roxygen
- aerobic
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