REVIEW PAPER

Citric Acid Cycle and Role of its Intermediates in Metabolism

Muhammad Akram

Published online: 26 September 2013

Springer Science+Business Media New York 2013

Abstract The citric acid cycle is the final common oxi-

dative pathway for carbohydrates, fats and amino acids. It

is the most important metabolic pathway for the energy

supply to the body. TCA is the most important central

pathway connecting almost all the individual metabolic

pathways. In this review article, introduction, regulation

and energetics of TCA cycle have been discussed. The

present study was carried out to review literature on TCA

cycle.

Keywords TCA cycle Energetics of TCA cycle Regulation of TCA cycle Amphibolic role of TCAcycle

Introduction

The citric acid cycle was proposed by Hans Adolf Krebs in

1937. The citric acid cycle is the primary metabolic pathway

for all the aerobic processes in an animal tissue. It is a series

of reactions that are important for the cells: C2 units or

acetyl-CoA that is derived from fats, carbohydrates and

lipids [12]. TCA cycle utilizes (indirectly) about 2/3 of the

total oxygen consumed by the body and generates about 2/3

of the total energy. The citric acid cycle is the final common

pathway for the oxidation of carbohydrate, protein and lip-

ids. TCA plays an important role in gluconeogenesis,

transamination, deamination and lipogenesis. Oxidation of

acetyl-CoA by TCA cycle accounts for 2/3rd of total oxygen

consumption and ATP production. Acetyl-CoA is obtained

from amino acids like leucine, tyrosine, isoleucine, lysine,

phenylalanine and tryptophan, triacylglycerol, carbohy-

drates and ketone bodies. In aerobic organisms the TCA is

amphibolic pathway, one that participates both in the cata-

bolic and anabolic processes [3]. Through its role in the

oxidative catabolism of carbohydrates, fatty acids and

amino acids, the cycle provides precursors for many bio-

synthetic pathways. Unlike the other metabolic pathways/

cycle, very few genetic abnormalities of TCA cycle are

known. This may be due to the vital importance of this

metabolic cycle for the survival of life. It has been observed

that stimulation of Krebs cycle activity in the presence of

ACTH may lead to increased production of steroid precur-

sor(s) of corticosterone which in turn creates an increased

demand for reducing equivalents for their conversion into

corticosterone.

History

Albert Szent-Gyorgyi and Hans Adolf Krebs [11] estab-

lished reactions and components of TCA in 1930.

Location of TCA cycle

The enzymes of TCA cycle are located in mitochondrial

matrix [8].

Enzymes of TCA Cycle [17]

Citrate synthase

Aconitase

Isocitrate dehydrogenase

Ketoglutarate dehydrogenase

M. Akram (&)Department of Eastern Medicine and Surgery, Faculty of

Medical and Health Sciences, The University of Poonch,

Azad Jammu and Kashmir, Pakistan

e-mail: makram_0451@hotmail.com; makram0451@gmail.com

123

Cell Biochem Biophys (2014) 68:475478

DOI 10.1007/s12013-013-9750-1

Succinyl-CoA synthase

Succinate dehydrogenase

Fumarase

Malate dehydrogenase

Sources of Acetyl-CoA

Acetyl-CoA is obtained from various sources including

carbohydrates in which glucose is broken down to pyruvic

acid and pyruvic acid is decarboxylated to acetyl-CoA.

Pyruvic acid is a three-carbon compound and acetyl-CoA is

a two-carbon compound. Acetyl-CoA (two-carbon com-

pound) is the starting point for the TCA cycle. Breakdown

of acetyl-CoA is the catabolic role of TCA cycle. Jones

et al. [9] have reported the sources of acetyl-CoA for

entering the tricarboxylic acid cycle as determined by

analysis of succinate 13C isotopomers.

Stages of Citric Acid Cycle

Step 1: Condensation

Acetyl-CoA (two-carbon compound) and oxaloacetate

(four-carbon compound) are converted to citrate (six-car-

bon compound). This reaction is catalyzed by citrate

synthetase.

Step 2: Isomerization of Citrate

Citrate is converted to cis-aconitate. This is catalyzed by

aconitase [6]. Cis-aconitate is an intermediate and is further

converted to isocitrate by aconitase. Aconitase is involved

in both reactions. In which first dehydration and then

rehydration occur and as a result final product isocitrate is

obtained

Step 3: Generation of CO2 by an NAD? Linked

Enzyme

Isocitrate is converted to alpha-ketoglutarate by isocitrate

dehydrogenase. Isocitrate is first dehydrogenated to

Oxalosuccinate which is an unstable compound and is

readily decarboxylated to alpha-ketoglutarate. In addition

to decarboxylation, this step produces a reduced nicotin-

amide adenine dinucleotide (NADH2) cofactor.

Step 4: A Second Oxidative Decarboxylation Step

This step is performed by a multi-enzyme complex, the

a-ketoglutarate dehydrogenation complex. In this reaction,

alpha-ketoglutarate is converted to succinyl-CoA by alpha-

ketoglutarate dehydrogenase (Guffon et al. 1993).

Step 5: Substrate-Level Phosphorylation

Succinyl-CoA is converted to succinate by succinic thio-

kinase. In this reaction GDP is converted to GTP

Step 6: Flavin-Dependent Dehydrogenation

Succinate is converted to fumaric acid by succinate dehy-

drogenase [5]. In this reaction, FAD is converted to

FADH2.

Step 7: Hydration of a CarbonCarbon Double Bond

Hydration of the C=C double bond occurs that is catalyzed

by Fumarate Hydratase (also known as Fumarase) and

malate is formed [2].

Step 8: A Dehydrogenation Reaction that Regenerates

Oxaloacetate

Malate is dehydrogenated to produce oxaloacetate by the

enzyme Malate Dehydrogenase. In this reaction NAD is

converted to NADH2. Oxaloacetate formed in this reaction

reacts with acetyl-CoA to form citrate in order to start

another round of the citric acid cycle [15].

Energetics of TCA Cycle [13]

3 NADH2 = 9 ATP

1 FADH2 = 2 ATP

1 GTP = 1 ATP

Total ATP = 12 ATP

Requirement of Oxygen by TCA cycle

There is no direct participation of oxygen in TCA. How-

ever, the cycle operates only under aerobic conditions [10].

Inhibitors of TCA

Fluoroacetate

It condenses with CoA and form fluoroacetyl-CoA. Fluo-

roacetyl-CoA condenses with oxaloacetate to form fluo-

rocitrate that inhibits aconitase, as a result citrate start to

accumulate. Fonnum et al. [7] have studied the use of

fluorocitrate and fluoroacetate in the brain metabolism.

476 Cell Biochem Biophys (2014) 68:475478

123

Arsenite

It inhibits alpha-ketoglutarate dehydrogenase

Malonate

It inhibits succinate dehydrogenase

Regulation of TCA

Three enzymes namely citrate synthase, isocitrate dehy-

drogenase and alpha-ketoglutarate dehydrogenase regulate

citric acid cycle. Wan et al. [16] have reported the regu-

lation of citric acid cycle by calcium.

Metabolic role of citric acid cycle

1. Oxidation of acetyl-CoA Glucose and fatty acids form

acetyl-CoA. It is converted to carbon di-oxide and

water. Abdel-aleem et al. [1] stated that fatty acid

oxidation is regulated by acetyl-CoA generated from

glucose oxidation that is evident in a study conducted

in isolated myocytes.

2. Integration with amino acids metabolism Many amino

acids after being deaminated give rise to compounds

that are also intermediate metabolites in citric acid

cycle [14].

3. Integration with lipid metabolism Fatty acids are also

first oxidized to acetyl-CoA which is then oxidized in

the citric acid cycle [4]

Biosynthetic function

Several intermediates of this cycle take part in the forma-

tion of substances of great physiological importance. Cit-

rate is provided to the lens of the eye, bone and the seminal

plasma. Citrate stimulates fatty acids synthesis by activat-

ing acetyl-CoA carboxylase

Conclusion

Citric acid cycle is one of the main metabolic pathways

that living cells utilize to completely oxidize biofuels to

carbon dioxide and water. The pyruvic acid formed as a

result of glycolysis is completely oxidized via this cycle.

Before pyruvic acid can enter citric acid cycle, it is con-

verted to acetyl-CoA. The acetyl-CoA thus produced is

then completely oxidized in citric acid cycle. This cycle not

only supplies energy but also provides many intermediates

required for the synthesis of amino acids, glucose, heme,

etc. The citric acid cycle is amphibolic, since it has other

metabolic roles in addition to oxidation. It takes place in

gluconeogenesis, transamination, deamination and the

synthesis of fatty acids.

References

1. Abdel-aleem, S., Nada, M. A., Sayed-Ahmed, M., Hendrickson,

S. C., St Louis, J., Walthall, H. P., et al. (1996). Regulation of

fatty acid oxidation by acetyl-CoA generated from glucose uti-

lization in isolated myocytes. Journal of Molecular and Cellular

Cardiology, 28(5), 825833.

2. Akiba, T., Hiraga, K., & Tuboi, S. (1984). Intracellular distri-

bution of fumarase in various animals. Journal of Biochemistry

(Tokyo), 96, 189195.

3. Baldwin, J. E., & Krebs, H. (1981). The evaluation of metabolic

cycle. Nature, 292, 291381.

4. Bowtell, J. L., Marwood, S., Bruce, M., Constantin-Teodosiu, D.,

& Greenhaff, P. L. (2007). Tricarboxylic acid cycle intermediate

pool size: Functional importance for oxidative metabolism in

exercising human skeletal muscle. Sports Medicine (Auckland, N.

Z.), 37(12), 10711088.

5. Briere, J. J., Favier, J., El Ghouzzi, V., Djouadi, F., Benit, P.,

Gimenez, A. P., et al. (2005). Succinate dehydrogenase defi-

ciency in human. Cellular and Molecular Life Sciences, 62,

23172324.

6. Costello, L. C., & Franklin, R. B. (1981). Aconitase activity,

citrate oxidation, and zinc inhibition in rat ventral prostate.

Enzyme, 26, 281287.

7. Fonnum, F., Johnsen, A., & Hassel, B. (1997). Use of fluoroci-

trate and fluoroacetate in the study of brain metabolism. Glia,

21(1), 106113.

8. Hellemond, J. J., Opperdoes, F. R., & Tielens, A. G. (2005). The

extraordinary mitochondrion and unusual citric acid cycle in

Trypanosoma brucei van. Biochemical Society Transactions,

33(5), 967971.

9. Jones, J. G., Sherry, A. D., Jeffrey, F. M., Storey, C. J., & Malloy,

C. R. (1993). Sources of acetyl-CoA entering the tricarboxylic

acid cycle as determined by analysis of succinate 13C isotopo-

mers. Biochemistry, 32(45), 1224012244.

10. Kamzolova, S. V., Shishkanova, N. V., Morgunov, I. G., & Fi-

nogenova, T. V. (2003). Oxygen requirements for growth and

citric acid production of Yarrowia lipolytica. FEMS Yeast

Research, 3(2), 217222.

11. Krebs, H. A. (1970). The history of the tricarboxylic acid cycle.

Perspectives in Biology and Medicine, 14(1), 154170.

12. Melendez, E., Waddell, T., & Cascante, M. (1996). The puzzle of

the citric acid cycle. Journal of Molecular Evolution, 43,

293303.

13. Nazaret, C., Heiske, M., Thurley, K., & Mazat, J. P. (2009).

Mitochondrial energetic metabolism: A simplified model of TCA

cycle with ATP production. Journal of Theoretical Biology,

258(3), 455464.

14. Reeds, P. J., Berthold, H. K., Boza, J. J., Burrin, D. G., Jahoor, F.,

Jaksic, T., et al. (1997). Integration of amino acid and carbon

intermediary metabolism: Studies with uniformly labeled tracers

and mass isotopomer analysis. European Journal of Pediatrics,

156(1), 5058.

Cell Biochem Biophys (2014) 68:475478 477

123

15. Srere, P. A. (1975). The enzymology of the formation and

breakdown of citrate. Advanced Enzymology, 43, 57.

16. Wan, B., LaNoue, K. F., Cheung, J. Y., & Scaduto, R. C, Jr.

(1989). Regulation of citric acid cycle by calcium. Journal of

Biological Chemistry, 264(23), 1343013439.

17. Zatta, P., Lain, E., & Cagnolini, C. (2000). Effects of aluminum

on activity of Krebs cycle enzymes and glutamate dehydrogenase

in rat brain homogenate. European Journal of Biochemistry,

267(10), 30493055.

478 Cell Biochem Biophys (2014) 68:475478

123

Citric Acid Cycle and Role of its Intermediates in MetabolismAbstractIntroductionHistoryLocation of TCA cycleEnzymes of TCA Cycle [17]

Sources of Acetyl-CoAStages of Citric Acid CycleStep 1: CondensationStep 2: Isomerization of CitrateStep 3: Generation of CO2 by an NAD+ Linked EnzymeStep 4: A Second Oxidative Decarboxylation StepStep 5: Substrate-Level PhosphorylationStep 6: Flavin-Dependent DehydrogenationStep 7: Hydration of a Carbon--Carbon Double BondStep 8: A Dehydrogenation Reaction that Regenerates Oxaloacetate

Energetics of TCA Cycle [13]Requirement of Oxygen by TCA cycle

Inhibitors of TCAFluoroacetateArseniteMalonate

Regulation of TCAMetabolic role of citric acid cycleBiosynthetic functionConclusionReferences