Chapter 16 Exam 3 Flashcards
What is cellular respiration?
cellular respiration = process by which the pyruvate produced by glycolysis is further oxidized to H2O and CO2
Stage 1: oxidation of fuels to acetyl-CoA
generates ATP, NADH, FADH2
Stage 2: oxidation of acetyl groups to CO2 in the citric acid cycle (tricarboxylic acid (TCA) cycle, Krebs cycle)
nearly universal pathway
generates NADH, FADH2, and one GTP
Stage 3: electron transfer chain and oxidative phosphorylation
generates the vast majority of ATP from catabolism
What is Coenzyme A (CoA-SH)?
coenzyme A has a reactive thiol (–SH) group that is critical to its role as an acyl carrier
the –SH group forms a thioester with acetate in acetyl-CoA
The acyl group attached to coenzyme A known as “activated” for group transfer.
What is mitochondrial pyruvate carrier (MPC)?
Pyruvate is Oxidized to Acetyl-CoA and CO2
mitochondrial pyruvate carrier (MPC) = an H+-coupled pyruvate-specific symporter in the inner mitochondrial membrane
What is pyruvate dehydrogenase (PDH) complex?
Pyruvate is Oxidized to Acetyl-CoA and CO2
pyruvate dehydrogenase (PDH) complex = highly ordered cluster of enzymes and cofactors that oxidizes pyruvate in the mitochondrial matrix to acetyl-CoA and CO2
–the series of chemical intermediates remain bound to the enzyme subunits
–regulation results in precisely regulated flux
What is oxidative decarboxylation?
oxidative decarboxylation = an irreversible oxidation process in which the carboxyl group is removed, forming CO2
(The PDH Complex Catalyzes an Oxidative Decarboxylation)
The PDH Complex Employs what Three Enzymes and Five Coenzymes to Oxidize Pyruvate?
three enzymes:
pyruvate dehydrogenase, E1
dihydrolipoyl transacetylase, E2
dihydrolipoyl dehydrogenase, E3
five coenzymes:
thiamine pyrophosphate (TPP)
lipoate
coenzyme A (CoA, CoA-SH)
flavin adenine dinucleotide (FAD)
nicotinamide adenine dinucleotide (NAD)
What is lipoate?
lipoate = coenzyme with two thiol groups that can undergo reversible oxidation to a disulfide bond (–S–S–)
serves as an electron (hydrogen) carrier and an acyl carrier
covalently linked to E2 via a lysine residue
What are the The PDH Complex Enzymes?
the PDH complex contains multiple copies of:
pyruvate dehydrogenase (E1)
dihydrolipoyl transacetylase (E2)
dihydrolipoyl dehydrogenase (E3)
an E2 core (of 24-60 copies) is surrounded by multiple and variable numbers of E1 and E3 copies
(The active site of E1 has noncovalently bound TPP. E2 has the prosthetic group lipoate, attached through an amide bond to the ε-amino group of a Lys residue.)
What is the E1-E2-E3 Structure of the PDH Complex?
the structure of the PDH complex is similar to other enzymes that catalyze oxidations:
α-ketoglutarate dehydrogenase
branched-chain α-keto acid dehydrogenase (breakdown of the branched-chain amino acids valine, isoleucine, and leucine)
in a given species, E3 is identical in all three complexes
similarities reflect a common evolutionary origin
they are paralogs
Why does the PDH Complex Channels its Intermediates through Five Reactions?
Leigh syndrome (subacute necrotizing encephalomyelopathy) is a rare, progressive, neurodegenerative disorder caused by defects in mitochondrial ATP production, primarily as a result of mutations in genes that encode proteins of the PDHC, the ETC, or ATP synthase. Both nuclear and mitochondrial DNA can be affected.
What are the first two steps of Oxidative Decarboxylation of Pyruvate?
pyruvate dehydrogenase, E1, with bound TPP catalyzes:
step 1: decarboxylation of pyruvate to the hydroethyl derivate
rate-limiting step
step 2: oxidation of the hydroethyl derivate to an acetyl group
electrons and the acetyl group are transferred from TPP to the lipoyllysyl group of E2
What are steps 3 of Oxidative Decarboxylation of Pyruvate?
dihydrolipoyl transacetylase, E2, catalyzes:
step 3: esterification of the acetyl moiety to one of the lipoyl –SH groups, followed by transesterification to CoA to form acetyl-CoA
What are steps 4 and 5 of Oxidative Decarboxylation of Pyruvate?
dihydrolipoyl dehydrogenase, E3, catalyzes:
step 4: electron transfer to regenerate the oxidized form of the lipoyllysyl group
step 5: electron transfer to regenerate the oxidized FAD cofactor, forming NADH
What is substrate channeling?
substrate channeling = the passage of intermediates from one enzyme directly to another enzyme without release
the long lipoyllysyl arm of E2 channels the substrate from the active site of E1 to E2 to E3
tethers intermediates to the enzyme complex
increases the efficiency of the overall reaction
minimizes side reactions
The Five-Reaction Sequence of the PDH Complex is an Example of Substrate Channeling
What are the reactions of the Citric Acid Cycle?
one oxaloacetate molecule can theoretically oxidize an infinite number of acetyl groups
energy from the four oxidations is conserved as NADH and FADH2
In Eukaryotes, why is the Mitochondrion the Site of Energy-Yielding Oxidative Reactions and ATP Synthesis?
isolated mitochondria contain all enzyme, coenzymes, and proteins needed for:
the citric acid cycle
electron transfer and ATP synthesis by oxidative phosphorylation
oxidation of fatty acids and amino acids to acetyl-CoA
oxidative degradation of amino acids to citric acid cycle intermediates
Why does the Sequence of Reactions in the Citric Acid Cycle Makes Chemical Sense?
complete oxidation of acetyl-CoA to CO2 extracts the maximum potential energy
direct oxidation to yield CO2 and CH4 is not biochemically feasible because organisms cannot oxidize CH4
carbonyl groups are more chemically reactive than a methylene group or methane
each step of the cycle involves either:
an energy-conserving oxidation
placing functional groups in position to facilitate oxidation or oxidative decarboxylation
How many steps does the Citric Acid cycle have? What does the cycle yield?
The Citric Acid Cycle has Eight Steps
-citrate formed from acetyl-CoA and oxaloacetate is oxidized to yield:
CO2
NADH
FADH2
GTP or ATP
Four- and five-carbon intermediates of the cycle serve as precursors for a wide variety of products.
What is citrate synthase?
citrate synthase = catalyzes the condensation of acetyl-CoA with oxaloacetate to form citrate
involves the formation of a transient intermediate, citroyl-CoA
large, negative ∆G′° is needed because [oxaloacetate] is normally very low
Formation of Citrate
What is the structure of Citrate Synthase?
binding of oxaloacetate creates a binding site for acetyl-CoA
induced fit decreases the likelihood of premature cleavage of the thioester bond of acetyl-CoA
What is the mechanism of Citrate Synthase?
What is aconitase (aconitate hydratase)?
(Formation of Isocitrate via Cis-Aconitate)
aconitase (aconitate hydratase) = catalyzes the reversible transformation of citrate to isocitrate through the intermediate cis-aconitate
addition of H2O to cis-aconitate is stereospecific
low [isocitrate] pulls the reaction forward
(Enzyme is inhibited by fluoroacetate, a rat poison.
Fluoroacetate forms fluoroacetyl CoA which condenses with oxaloacetate to form fluorocitrate – a potent inhibitor of aconitase. This results in citrate accumulation.)
What is α-ketoglutarate dehydrogenase complex?
α-ketoglutarate dehydrogenase complex = catalyzes the oxidative decarboxylation of α-ketoglutarate to succinyl-CoA and CO2
energy of oxidation is conserved in the thioester bond of succinyl-CoA
(Oxidation of α-Ketoglutarate to Succinyl-CoA and CO2)
Why is the Mechanism for Oxidative Decarboxylation conserved?
pathways use the same five cofactors, similar multienzyme complexes, and the same enzymatic mechanism
have homologous E1 and E2 and identical E3
example of gene duplication and divergent evolution
What is succinyl-CoA synthetase (succinic thiokinase)?
(Conversion of Succinyl-CoA to Succinate)
succinyl-CoA synthetase (succinic thiokinase) = catalyzes the breakage of the thioester bond of succinyl-CoA to form succinate
energy released drives the synthesis of a phosphoanhydride bond in either GTP or ATP (substrate level phosphorylation)
Animal cells have two isozymes of succinyl-CoA synthetase, one specific for ADP and the other for GDP.
What is the The Succinyl-CoA Synthetase Reaction (1 of 2)?
enzyme molecule becomes phosphorylated at a His residue in the active site
phosphoryl group is then transferred to ADP or GDP to form ATP or GTP
animal cells have specific isozymes for ADP and GDP
What is The Succinyl-CoA Synthetase Reaction (2 of 2)?
power helices place the partial positive charges of the helix dipole near the phosphate group of the α chain phosphorylated His246 to stabilize the phosphoenzyme intermediate
What is succinate dehydrogenase?
succinate dehydrogenase = flavoprotein that catalyzes the reversible oxidation of succinate to fumarate
integral protein of the mitochondrial inner membrane in eukaryotes
contains three iron-sulfur clusters and covalently bound FAD
(Oxidation of Succinate to Fumarate)
succinate dehydrogenase is an integral protein of the mitochondrial inner membrane; in bacteria, of the plasma membrane.
What is Malonate a competitive inhibitor for?
Malonate is a Strong Competitive Inhibitor of Succinate Dehydrogenase
malonate = an analog of succinate
not normally present in cells
addition to mitochondria in-vitro blocks citric acid cycle activity
What is fumarase?
fumarase = catalyzes the reversible hydration of fumarate to L-malate
transition state is a carbanion
(Hydration of Fumarate to Malate)
What is L-malate dehydrogenase?
L-malate dehydrogenase = catalyzes the oxidation of L-malate to oxaloacetate, coupled to the reduction of NAD+
low [oxaloacetate] pulls the reaction forward
regenerates oxaloacetate for citrate synthesis
(Oxidation of Malate to Oxaloacetate)
How is energy of Oxidations in the cycle efficiently conserved?
energy released by oxidation is conserved in the production of:
3 NADH
1 FADH2
1 GTP (or ATP)
What are Anaplerotic Reactions?
when intermediates are shunted from the citric acid cycle to other pathways, they are replenished
anaplerotic reactions = chemical reactions that replenish intermediates
(Anaplerotic Reactions Replenish Citric Acid Cycle Intermediates)
What is the role of Biotin in the Pyruvate Carboxylase reaction?
the two steps in carboxylation of pyruvate occur at separate active sites
the arm of biotin transfers activated carboxyl groups from the first active site to the second
(Pyruvate carboxylase has four identical subunits, each containing a molecule of biotin covalently attached through an amide linkage to theε-amino group of a specific Lys residue in the enzyme active site)
What do biological tethers allow?
Biological Tethers Allow Flexibility
all participate in substrate channeling through the flexible tethers that move intermediates from one active site to the next
Review the Regulation of Metabolite Flow Through the Citric Acid Cycle
What is PDH kinase?
PDH kinase = inhibits the PDH complex by phosphorylation
allosterically activated by products of the complex
inhibited by substrates of the complex
Covalent Modification of the PDH Complex
What is PDH phosphatase?
PDH phosphatase = reverses the inhibition by PDH kinase
Covalent Modification of the PDH Complex
What can Mutations in Citric Acid Cycle Enzymes cause?
mutations in succinate dehydrogenase and fumarase cause tumors, defining them as tumor suppressors
many glial cell tumors have mutant NADPH-dependent isocitrate dehydrogenase
lose ability to convert isocitrate to α-ketoglutarate
gain ability to convert α-ketoglutarate to 2-hydroxyglutarate
