Glyoxylate Bypass Operon of Escherichia Coli: Cloning and Determination of the Functional Map

Overview

Journal J Bacteriol

Publisher American Society for Microbiology

Specialty Microbiology

Date 1988 Jan 1

PMID 2826399

Citations 32

Authors

T Chung

D J Klumpp

D C LaPorte

Affiliations

Soon will be listed here.

Abstract

In Escherichia coli, a single operon encodes the metabolic and regulatory enzymes of the glyoxylate bypass. The metabolic enzymes, isocitrate lyase and malate synthase, are expressed from aceA and aceB, and the regulatory enzyme, isocitrate dehydrogenase kinase/phosphatase, is expressed from aceK. We cloned this operon and determined its functional map by deletion analysis. The order of the genes in this operon is aceB-aceA-aceK, with aceB proximal to the promoter, consistent with the results of previous experiments using genetic techniques. The promoter was identified by S1 nuclease mapping, and its nucleotide sequence was determined. Isocitrate lyase and malate synthase were readily identified by autoradiography after the products of the operon clone were labeled by the maxicell procedure and then resolved by electrophoresis. In contrast, isocitrate dehydrogenase kinase/phosphatase, expressed from the same plasmid, was undetectable. This observation is consistent with a striking downshift in expression between aceA and aceK.

Citing Articles

Malate synthase contributes to the survival of Salmonella Typhimurium against nutrient and oxidative stress conditions.

Sarkhel R, Apoorva S, Priyadarsini S, Sridhar H, Bhure S, Mahawar M Sci Rep. 2022; 12(1):15979.

PMID: 36155623 PMC: 9510125. DOI: 10.1038/s41598-022-20245-0.

Microbial Upgrading of Acetate into Value-Added Products-Examining Microbial Diversity, Bioenergetic Constraints and Metabolic Engineering Approaches.

Kutscha R, Pflugl S Int J Mol Sci. 2020; 21(22).

PMID: 33233586 PMC: 7699770. DOI: 10.3390/ijms21228777.

Metabolic Feedback Inhibition Influences Metabolite Secretion by the Human Gut Symbiont Bacteroides thetaiotaomicron.

Catlett J, Catazaro J, Cashman M, Carr S, Powers R, Cohen M mSystems. 2020; 5(5).

PMID: 32873608 PMC: 7470985. DOI: 10.1128/mSystems.00252-20.

Truncation of C-Terminal Intrinsically Disordered Region of Mycobacterial Rv1915 Facilitates Production of "Difficult-to-Purify" Recombinant Drug Target.

Antil M, Gouin S, Gupta V Front Bioeng Biotechnol. 2020; 8:522.

PMID: 32548107 PMC: 7273500. DOI: 10.3389/fbioe.2020.00522.

Differential Metabolism of a Two-Carbon Substrate by Members of the Genus.

Baeza L, da Mata F, Pigosso L, Pereira M, de Souza G, Coelho A Front Microbiol. 2017; 8:2308.

PMID: 29230201 PMC: 5711815. DOI: 10.3389/fmicb.2017.02308.

References

Garnak M, Reeves H . Purification and properties of phosphorylated isocitrate dehydrogenase of Escherichia coli. J Biol Chem. 1979; 254(16):7915-20. View

Garnak M, Reeves H . Phosphorylation of Isocitrate dehydrogenase of Escherichia coli. Science. 1979; 203(4385):1111-2. DOI: 10.1126/science.34215. View

Maloy S, Bohlander M, Nunn W . Elevated levels of glyoxylate shunt enzymes in Escherichia coli strains constitutive for fatty acid degradation. J Bacteriol. 1980; 143(2):720-5. PMC: 294349. DOI: 10.1128/jb.143.2.720-725.1980. View

Maloy S, Nunn W . Genetic regulation of the glyoxylate shunt in Escherichia coli K-12. J Bacteriol. 1982; 149(1):173-80. PMC: 216607. DOI: 10.1128/jb.149.1.173-180.1982. View

LaPorte D, Koshland Jr D . A protein with kinase and phosphatase activities involved in regulation of tricarboxylic acid cycle. Nature. 1982; 300(5891):458-60. DOI: 10.1038/300458a0. View

Wang J, Koshland Jr D . The reversible phosphorylation of isocitrate dehydrogenase of Salmonella typhimurium. Arch Biochem Biophys. 1982; 218(1):59-67. DOI: 10.1016/0003-9861(82)90321-6. View

Burton Z, Gross C, Watanabe K, Burgess R . The operon that encodes the sigma subunit of RNA polymerase also encodes ribosomal protein S21 and DNA primase in E. coli K12. Cell. 1983; 32(2):335-49. DOI: 10.1016/0092-8674(83)90453-1. View

HAWLEY D, McClure W . Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res. 1983; 11(8):2237-55. PMC: 325881. DOI: 10.1093/nar/11.8.2237. View

LaPorte D, Walsh K, Koshland Jr D . The branch point effect. Ultrasensitivity and subsensitivity to metabolic control. J Biol Chem. 1984; 259(22):14068-75. View

10.

Belasco J, Beatty J, ADAMS C, von Gabain A, Cohen S . Differential expression of photosynthesis genes in R. capsulata results from segmental differences in stability within the polycistronic rxcA transcript. Cell. 1985; 40(1):171-81. DOI: 10.1016/0092-8674(85)90320-4. View

11.

LaPorte D, Thorsness P, Koshland Jr D . Compensatory phosphorylation of isocitrate dehydrogenase. A mechanism for adaptation to the intracellular environment. J Biol Chem. 1985; 260(19):10563-8. View

12.

LaPorte D, Chung T . A single gene codes for the kinase and phosphatase which regulate isocitrate dehydrogenase. J Biol Chem. 1985; 260(28):15291-7. View

13.

Mackintosh C, Duncan K, HOLMS W, Nimmo H . Molecular cloning and over-expression of the glyoxylate bypass operon from Escherichia coli ML308. Biochem J. 1987; 242(3):661-5. PMC: 1147762. DOI: 10.1042/bj2420661. View

14.

LOWRY O, ROSEBROUGH N, FARR A, RANDALL R . Protein measurement with the Folin phenol reagent. J Biol Chem. 1951; 193(1):265-75. View

15.

Kornberg H, MADSEN N . Synthesis of C4-dicarboxylic acids from acetate by a glyoxylate bypass of the tricarboxylic acid cycle. Biochim Biophys Acta. 1957; 24(3):651-3. DOI: 10.1016/0006-3002(57)90268-8. View

16.

Kornberg H . The role and control of the glyoxylate cycle in Escherichia coli. Biochem J. 1966; 99(1):1-11. PMC: 1264949. DOI: 10.1042/bj0990001. View

17.

Laemmli U . Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970; 227(5259):680-5. DOI: 10.1038/227680a0. View

18.

LaPorte D, Koshland Jr D . Phosphorylation of isocitrate dehydrogenase as a demonstration of enhanced sensitivity in covalent regulation. Nature. 1983; 305(5932):286-90. DOI: 10.1038/305286a0. View

19.

Nimmo G, Nimmo H . The regulatory properties of isocitrate dehydrogenase kinase and isocitrate dehydrogenase phosphatase from Escherichia coli ML308 and the roles of these activities in the control of isocitrate dehydrogenase. Eur J Biochem. 1984; 141(2):409-14. DOI: 10.1111/j.1432-1033.1984.tb08206.x. View

20.

Borthwick A, HOLMS W, Nimmo H . The phosphorylation of Escherichia coli isocitrate dehydrogenase in intact cells. Biochem J. 1984; 222(3):797-804. PMC: 1144244. DOI: 10.1042/bj2220797. View