Article: Detailed enzyme kinetics in terms of biochemical species: study of citrate synthase

The compulsory-ordered ternary catalytic mechanism for two-substrate two-product enzymes is analyzed to account for binding of inhibitors to each of the four enzyme states and to maintain the relationship between the kinetic constants and the reaction equilibrium constant. The developed quasi-steady flux expression is applied to the analysis of data from citrate synthase to determine and parameterize a kinetic scheme in terms of biochemical species, in which the effects of pH, ionic strength, and cation binding to biochemical species are explicitly accounted for in the analysis of the data. This analysis provides a mechanistic model that is consistent with the data that have been used support competing hypotheses regarding the catalytic mechanism of this enzyme.

Citing Articles

Human citrate synthase kinetic simulation to fit rapid, direct, and thiol probe coupled kinetic data.

Shackelford N, Zavodny Z, Schindler S, Fancher N, Thomas A, Moxley M Biochem Biophys Rep. 2025; 41:101914.

PMID: 39886071 PMC: 11780148. DOI: 10.1016/j.bbrep.2025.101914.

CitA activity is modulated by cysteine oxidation and pyruvate binding.

Pathirage R, Favrot L, Petit C, Yamsek M, Singh S, Mallareddy J RSC Med Chem. 2023; 14(5):921-933.

PMID: 37252106 PMC: 10211323. DOI: 10.1039/d3md00058c.

The Expanding Computational Toolbox for Engineering Microbial Phenotypes at the Genome Scale.

Zielinski D, Patel A, Palsson B Microorganisms. 2020; 8(12).

PMID: 33371386 PMC: 7767376. DOI: 10.3390/microorganisms8122050.

Temperature-Dependent Estimation of Gibbs Energies Using an Updated Group-Contribution Method.

Du B, Zhang Z, Grubner S, Yurkovich J, Palsson B, Zielinski D Biophys J. 2018; 114(11):2691-2702.

PMID: 29874618 PMC: 6129446. DOI: 10.1016/j.bpj.2018.04.030.

Determination of the catalytic mechanism for mitochondrial malate dehydrogenase.

Dasika S, Vinnakota K, Beard D Biophys J. 2015; 108(2):408-19.

PMID: 25606688 PMC: 4302198. DOI: 10.1016/j.bpj.2014.11.3467.

References

1.

Frieden C, ALBERTY R . The effect of pH on fumarase activity in acetate buffer. J Biol Chem. 1955; 212(2):859-68. View

2.

Teusink B, Passarge J, Reijenga C, Esgalhado E, van der Weijden C, Schepper M . Can yeast glycolysis be understood in terms of in vitro kinetics of the constituent enzymes? Testing biochemistry. Eur J Biochem. 2000; 267(17):5313-29. DOI: 10.1046/j.1432-1327.2000.01527.x. View

3.

Lambeth M, Kushmerick M . A computational model for glycogenolysis in skeletal muscle. Ann Biomed Eng. 2002; 30(6):808-27. DOI: 10.1114/1.1492813. View

4.

Smith C, Williamson J . Inhibition of citrate synthase by succinyl-CoA and other metabolites. FEBS Lett. 1971; 18(1):35-38. DOI: 10.1016/0014-5793(71)80400-3. View

5.

Mulquiney P, Kuchel P . Model of 2,3-bisphosphoglycerate metabolism in the human erythrocyte based on detailed enzyme kinetic equations: equations and parameter refinement. Biochem J. 1999; 342 Pt 3:581-96. PMC: 1220499. View

6.

Matsuoka Y, SRERE P . Kinetic studies of citrate synthase from rat kidney and rat brain. J Biol Chem. 1973; 248(23):8022-30. View

7.

Alberty R . Relations between biochemical thermodynamics and biochemical kinetics. Biophys Chem. 2006; 124(1):11-7. DOI: 10.1016/j.bpc.2006.05.024. View

8.

Vinnakota K, Kemp M, Kushmerick M . Dynamics of muscle glycogenolysis modeled with pH time course computation and pH-dependent reaction equilibria and enzyme kinetics. Biophys J. 2006; 91(4):1264-87. PMC: 1518652. DOI: 10.1529/biophysj.105.073296. View

9.

SRERE P . Citrate-condensing enzyme-oxalacetate binary complex. Studies on its physical and chemical properties. J Biol Chem. 1966; 241(9):2157-65. View

10.

Shepherd D, Garland P . The kinetic properties of citrate synthase from rat liver mitochondria. Biochem J. 1969; 114(3):597-610. PMC: 1184933. DOI: 10.1042/bj1140597. View

11.

Jamshidi N, Edwards J, Fahland T, Church G, Palsson B . Dynamic simulation of the human red blood cell metabolic network. Bioinformatics. 2001; 17(3):286-7. DOI: 10.1093/bioinformatics/17.3.286. View

12.

ALBERTY R . Levels of thermodynamic treatment of biochemical reaction systems. Biophys J. 1993; 65(3):1243-54. PMC: 1225844. DOI: 10.1016/S0006-3495(93)81146-9. View

Detailed Enzyme Kinetics in Terms of Biochemical Species: Study of Citrate Synthase