ME03 - TCA Flashcards
CELLULAR RESPIRATION
GLYCOLYSIS – CAC – ETC – OxPhos
What is TCA
Because the cycle begins with the formation of citric acid which is a carboxylic acid with 3 COOH grps ! also called Tricarboxylic acid cycle
TCA occurs only in the presence of O2
Since it is an aerobic process
Where does it takes place
takes place in the mitochondrial matrix,except
succinate dehydrogenase (inner membrane)
Processes involved in TCA
involves oxidation of pyruvic acid into CO2 and H20 final common pathway for the oxidation of CHO, CHON, fats
Functions of TCA
_ Provides majority of ATP for energy
_ Interconverts amino acids (transamination-
deamination)-creates amino acids or destroys
amino acids (producing glucose)
_ Has a crucial role in fatty acid a synthesis
Biomedical Importance of TCA
CAC has a central role in gluconeogenesis, lipogenesis,
and interconversion of AA
_ Liver is the only tissue in which al loccur to a significant extent Hepatitis (large no of hepatic cells are damaged) and Cirrhosis (hepatic cells are replaced with connective tissue) pose repercussions in these processes.
Genetic defects of CAC enzyme
_ Associatedwithneurologicaldamageasaresultof
impaired ATP formation in the CNS
Steps in TCA
series of 9 enzyme controlled reactions!ultimate goal
is to produce ENERGY (ATP)
reactions of the CAC liberate reducing equivalents and
CO2
Sources of Energy in TCA
Beta-oxidation of FAs and Pyruvates from
glucose
Formation of Acetyl CoA: Oxidative Decarboxylation
Enzyme: Pyruvate Dehydrogenase
3 C pyruvate ! 2-C Acetyl CoA
_ 2H+ and one CO2 molecule are removed :Oxidative decarboxylation
_ 2H+areacceptedbyNADandNADisconverted (REDUCED) into NADH
Substrates and Products in TCA
Substrates: Acetyl CoA and Oxaloacetate
Products: 12 ATP (from 3 NADH, 1 FADH, 1 GTP) , 2CO2, H2O,heat (inefficient waste product)
Enzymes in TCA
Can ! CITRATE
I ! ISOCITRATE
Ask ! ALPHAKETOGLUTARATE
Some ! SUCCINYL CoA
Special ! SUCCINATE (Succinyl CoA come first coz Succinate is always late)
Friendly ! FUMARATE
Moment ! MALATE
Overnight? ! OXALOACETATE
- Acetyl CoA + Oxaloacetate»_space; CITRATE
Enzyme: Citrate Synthase
Acetyl CoA (2-C) , OAA (4-C), Citrate (6-C)
Only a small amt of OAA is needed for the oxidation of a
large qty of Acetyl CoA ! it can be considered as playing
catalytic role.
C-C bond formation b/w methyl carbon of Acetyl CoA
(CH3- CO-S-CoA) and carbonyl carbon of OAA
The thioester bond of citryl CoA is hydrolyzed releasing
citrate and CoASH!an exothermic rxn
- Citrate»_space; ISOCITRATE
Enzyme: Aconitase (isomerization)
The reaction occurs in two steps:
_ Dehydration to cis-aconitate
_ Rehydration to isocitrate
Although citrate is a symmetric molecule, aconitase
reacts with citrate asymmetrically, so that 2-C atoms that were lost in subsequent reactions of the cycle are not those that were added from acetyl CoA.
_ Thisasymmetricbehavioristheresultofchanneling -transfer of product of citrate synthase directly onto the active site of aconitase, without entering free soln.
_ ItprovidesintegrationofCACactivityandtheprovision of citrate in the cytosol as a source of acetyl CoA for FA syn.
_ Inhibits aconitase
_ Toxic because fluoroacetyl CoA condenses with OAA
to form fluorocitrate causing citrate to accumulate
FLUROACETATE
3&4. Oxidative Decarboxylation
Decarboxylation:
6 C Isocitrate!5 C a-ketoglutarate
5-C a-ketoglutarate!4 C Succinyl CoA
Oxidation (2H are removed, 2H are accepted by NAD and NAD is converted to NADH+ and H+)
Isocitrate» 5 C a-ketoglutarate _Dehydration rxn:Isocitrate to Oxalosuccinateby isocitrate dehydrogenase
_ Thenundergoesdecarboxylationto
alphaketoglutarate.
_ DecarboxylationrequiresMg++orMn++ions
3 isozymes of isocitrate dehydrogenase:
i. NAD+ found only in the mitochondria
ii. Other two uses NADP+ and are found in the
mitochondria and cytosol
Respiratory-chain-linked oxidation of isocitrate proceeds almost completely through _____________
the NAD+ dependent enzyme
Alphaketoglutarate»_space; Succinyl CoA
Alpha ketoglutarate undergoes oxidative decarboxylation Catalyzed by a multi-enzyme complex similar to that involved in the oxidative decarboxylation of pyruvate.
The alphaketoglutarate dehydrogenase complex requires
the same cofactors as the pyruvate dehydrogenase complex thiamine diphosphate, lipoate, NAD+, FAD, CoA.
The equilibrium of this rxn is so much in favor of Succinyl CoA formation that it must be considered to be physiologically unidirectional.
In pyruvate oxidation,this inhibits alphaketoglutarate dehydrogenase complex_ Alphaketoglutarateaccumulates
ARSENITE
- Succinyl CoA»Succinate
Enzyme: Succinyl CoA Synthase (Succinate thiokinase) Yields GTP which fuels the conversion of ADP to ATP
ONLY example in the CAC of substrate level phosphorylation
In Succinyl CoA»Succinate, Tissues in w/c gluconeogenesis occurs (liver and kidney) contain two isozymes of succinate thiokinase.
_ One specific for GTP
_ The other, specific for ADP.
GTP formed is used for
DecarboxylationofOAAtoPEP(Phosphoenol
pyruvate) in gluconeogenesis.
_ Providesaregulatorylinkb/wCACactivity _ Withdrawal of OAA for gluconeogenesis
Metabolism of Ketone bodies in extrahepatic tissues involve
transfer of CoA from Succinyl CoA to acetoacetate forming acetoacetylCoA
__Enzyme: Succinyl CoA Acetoacetate- CoA transferase (thioophorase)
Have only the isoenzyme that uses ADP
Nongluconeogenic tissues
- Oxidation
Succinate!Fumarate by the removal of 2 H+ atoms
Enzyme: Succinate Dehydrogenase
_ The enzyme contains FAD and Fe-S protein and directly reduces ubiquinone in the ETC.
2H+ atoms are accepted by FAD and it forms FADH2
Succinate dehydrogenase (electron carrier II) is a direct
part of the ETC!bound in the inner mitochondrial membrane.
- Hydration
Fumarate ! Malate
_ Catalyzestheaddnofwateracrossthedoublebond
Enzyme: Fumarase
of the fumarate.
- DEHYDROGENATION - THE FINAL STEP
Malate (4-C)!Oxaloacetate (4-C)
Enzyme: Malate Dehydrogenase
2H+ removed!accepted by NAD!NADH + H+
A rxn requiring NAD+
Although the equilibrium of this rxn strongly favors
malate, the net flux is to OAA because of the continual removal of OAA (to form citrate, as a substrate for gluconeogenesis, or to undergo transamination to aspartate) and also the continual reoxidation of NADH.
Acetyl CoA vs Pyruvate
ATPs from substrate level phosphorylation: Both 1 for ACoa and Pyruvate ATPs from NADH: 9 for Acoa 12 for Pyruvate | ATPS from FADH2 - both 2ATPs for each Total Yield ATP: 12 for ACoa and 15 for Pyruvate
Steps produce ATP uses oxidative phosphorylation mainly except for
Synthesis of Succinate
Used in gluconeogenic step oxaloactetate to PEP
GTP
Yields in ONE Turn of Krebs Cycle
2-C molecule (Acetyl CoA) enters the cycle &joins a 4-C molecule (Oxaloacetate) ! Citric Acid
In series of steps, H+ and high energy electrons are removed from the parent molecule!Oxidation
Three NAD+ are converted into 3 NADH & 3H+
(transferred to the ETC, yield ~2.5 ATP)
One FAD is converted into 1 FADH2 (transferred to the
ETC, yield ~1.5 ATP)
One GTP (ATP) is made (by substrate level
phosphorylation!succinate thiokinase)
Two CO2 are released
At the end of the cycle!nothing remains of the original
glucose molecule .
5 Vitamin B tickets
_ Riboflavin (B1): in the form of FAD a cofactor in succinate dehydrogenase.
_ Niacin(B2):intheformofNADtheelectronacceptor for Isocitrate DH, alphaketoglutarate DH, and malate DH
_ Thiamin:asthiamindiphosphateaspartofCoAthe coenzyme for decarboxylation in the alphaketoglutarate DH
_ Pantothenicacid:aspartofCoAthecofactor attached to active carboxylic acid resisdues such as acetyl-CoA and Succinyl-CoA
_ LipoicAcid(B4)
CAC is also a pathway for interconversion of metabolites and A pathway for providing substrates for:
TransaminationanddeaminationofAA and AASynthesisbytransamination
Role in Bioenergetics-ATP Yield from TCA
_ Forgluconeogenesis
_ AndforFAsynthesis
The CAC is an amphibolic pathway
_ Becauseitparticipatesinbothcatabolism(Glycolysis, Beta Oxidation of triglycerides) and anabolism (protein/ lipid synthesis, glycogenesis)
Anaplerotic Reactions
_ Reactionsthatformintermediates(sources)of
components of the TCA cycle in order to replenish them especially during the scarcity of inhouse intermediates.
TCA - ATP Yield from TCA
Examples:
_ Aspartate ! OAA
_ Glutamate ! Alphaketoglutarate
_ FFAs!Succinyl CoA
_ Adenylsuccinate ! Fumarate
The CAC Takes Part in Gluconeogenesis, Transamination, and Deamination
All intermediates of CAC are glucogenic, since they can give rise to OAA and hence net production of glucose (in the liver and kidney, the organs that carry out gluconeogenesis).
Transamination Reactions: TCA intermediates to AA
Deamination Reactions: AA to intermediates
Enzyme: Phosphoenolpyruvate Key enzyme in catalyzing net transfer out of the cycle into GLUCONEOGENESIS
Catalyzes decarboxylation of OAA to PEP with GTP acting as donor
Formation of OAA from Pyruvate»_space;one example of anaplerotic rxn.
Enzyme: Pyruvate carboxylase
Impt in maintaining an adequate concentration of OAA for the condensation reaction with acetyl CoA If Acetyl CoA accumulates, it acts as both:
Allosteric activator of pyruvate carboxylase Inhibitor of Pyruvate dehydrogenase!Thereby ensuring a supply of OAA
Lactate enters the cycle via the oxidation of pyruvate and then carboxylation to OAA.
Amino transferase (transaminase) rxns form:
Pyruvate from Alanine
OAA from aspartate
Alphaketoglutarate from Glutamate
These reactions are reversible , CAC serves as a source of Carbon skeletons for the synthesis of these AA.
Other AA contribute to gluconeogenesis because the carbon skeletons give rise to CAC intermediates
Alanine, Cysteine, Glycine, Hydroxyproline, Serine,
Threonine, and Tryptophan yield PYRUVATE.
Arginine, Histidine, Glutamine, and Proline yield
ALPHAKETOGLUTARATE
Isoleucine,Methionine, and Valine yield SUCCINYL
COA
Tyrosine and Phenylalanine yield FUMARATE
In Ruminants PROPRIONATE (major glucogenic
prod of rumen fermentation) yield SUCCINYL COA via the methylmalonyl pathway.
CAC Takes Part in FA Synthesis
Acetyl CoA formed from Pyruvate by the action of Pyruvate Dehydrogenase!is the major substrate for long chain FFA
Pyruvate Dehydrogenase is a mitochondrial enzyme and FA synthesis is a cytosolic pathway
Acetyl CoA is made available in the cytosol from CITRATE synthesized in the mitochondria then transported to the cytosol and cleaved in a rxn catalyzed by ATP CITRATE LYASE.
CITRATE is only available for transport out of the mitochondria when ACONITASE is SATURATED with substrate, and citrate cannot be channeled directly from citrate synthase onto aconitase.
This ensures that CITRATE is used for FA SYN only when there is an adequate amount to ensure continued activity of the cycle.
CITRATE transports ACETYL COA from the mitochondrial matrix and into the cytoplasm to initiate fatty acid synthesis (Citrate Shuttle)
Other functions
Succinyl CoA can be used for heme synthesis and to activate ketone bodies in extra hepatic tissues
Malate can be used for gluconeogenesis
Regulation of CAC Depends Primarily on a Supply of Oxidized Cofactors
CAC is an energy yielding metabolism and RESPIRATORY CONTROL via the respiratory chain and oxidative phosphorylation regulates the CAC. CAC is immediately dependent on the supply of NAD+, which in turn, because of tight coupling b/w oxidation and phosphorylation, is dependent on the availability of ADP and hence, ultimately on the rate of utilization of ATP in chemical and physical work.
Individual enzymes of the CAC are regulated.
Pyruvate dehydrogenase
Citrate Synthase
Isocitrate dehydrogenase
Alphaketoglutarate dehydrogenase
What happens when Dehydrogenases are activated by Ca++
increases in concentration in muscular contraction and secretion, when there is increased energy demand.
In the brain!largely dependent on CHO to supply acetyl CoA!control of CAC may occur at pyruvate dehydrogenase.
Enzymes are responsive to energy status as shown by [ATP]/[ADP] AND [NADH]/[NAD] ratios.
Allosteric inhibition of CITRATE SYNTHASE by ATP and long chain fatty acyl-CoA
Allosteric activation of mitochondrial NAD-dependent ISOCITRATE DEHYDROGENASE by ADP is counteracted by ATP and NADPH
The ALPHAKETOGLUTARATE DH COMPLEX is regulated the same way as Pyruvate dehydrogenase.
What happens when SUCCINATE DH is inhibited by OAA
OAA, as controlled by the malate DH, depends on the [NADH]/[NAD+] ratio.
The Km for OAA of citrate synthase is of the same order of magnitude as the intramitochondrial conc, it is likely that the conc. of OAA controls the rate of CITRATE formation.
TRUE
Inhibitors of TCA
Fluoroacetate | Arsenite | Malonate
Which step is rate-limiting?
Isocitrate - a-ketoglutarate
Which steps have biochemical significance?
NADH, FADH, GTP
