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Isocitrate dehydrogenase kinase/phosphatase: aceK alleles that express kinase but not phosphatase activity.
J Bacteriol. 1991 Mar; 173(5):1801-6.JB

Abstract

For Escherichia coli, growth on acetate requires the induction of the enzymes of the glyoxylate bypass, isocitrate lyase and malate synthase. The branch point between the glyoxylate bypass and the Krebs cycle is controlled by phosphorylation of isocitrate dehydrogenase (IDH), inhibiting that enzyme's activity and thus forcing isocitrate through the bypass. This phosphorylation cycle is catalyzed by a bifunctional enzyme, IDH kinase/phosphatase, which is encoded by aceK. We have employed random mutagenesis to isolate novel alleles of aceK. These alleles were detected by the loss of ability to complement an aceK null mutation. The products of one class of these alleles retain IDH kinase activity but have suffered reductions in IDH phosphatase activity by factors of 200 to 400. Selective loss of the phosphatase activity also appears to have occurred in vivo, since cells expressing these alleles exhibit phenotypes which are reminiscent of strains lacking IDH; these strains are auxotrophic for glutamate. Assays of cell-free extracts confirmed that this phenotype resulted from nearly quantitative phosphorylation of IDH. The availability of these novel alleles of aceK allowed us to assess the significance of the precise control which is a characteristic of the IDH phosphorylation cycle in vivo. The fractional phosphorylation of IDH was varied by controlled expression of one of the mutant alleles, aceK3, in a wild-type strain. Reduction of IDH activity to 50% of the wild-type level did not adversely affect growth on acetate. However, further reductions inhibited growth, and growth arrest occurred when the IDH activity fell to 15% of the wild-type level. Thus, although wild-type cells maintain a precise effective IDH activity during growth on acetate, this precision is not critical.

Authors+Show Affiliations

Department of Biochemistry, University of Minnesota, Minneapolis 55455.No affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

1847910

Citation

Ikeda, T, and D C. LaPorte. "Isocitrate Dehydrogenase Kinase/phosphatase: aceK Alleles That Express Kinase but Not Phosphatase Activity." Journal of Bacteriology, vol. 173, no. 5, 1991, pp. 1801-6.
Ikeda T, LaPorte DC. Isocitrate dehydrogenase kinase/phosphatase: aceK alleles that express kinase but not phosphatase activity. J Bacteriol. 1991;173(5):1801-6.
Ikeda, T., & LaPorte, D. C. (1991). Isocitrate dehydrogenase kinase/phosphatase: aceK alleles that express kinase but not phosphatase activity. Journal of Bacteriology, 173(5), 1801-6.
Ikeda T, LaPorte DC. Isocitrate Dehydrogenase Kinase/phosphatase: aceK Alleles That Express Kinase but Not Phosphatase Activity. J Bacteriol. 1991;173(5):1801-6. PubMed PMID: 1847910.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Isocitrate dehydrogenase kinase/phosphatase: aceK alleles that express kinase but not phosphatase activity. AU - Ikeda,T, AU - LaPorte,D C, PY - 1991/3/1/pubmed PY - 1991/3/1/medline PY - 1991/3/1/entrez SP - 1801 EP - 6 JF - Journal of bacteriology JO - J Bacteriol VL - 173 IS - 5 N2 - For Escherichia coli, growth on acetate requires the induction of the enzymes of the glyoxylate bypass, isocitrate lyase and malate synthase. The branch point between the glyoxylate bypass and the Krebs cycle is controlled by phosphorylation of isocitrate dehydrogenase (IDH), inhibiting that enzyme's activity and thus forcing isocitrate through the bypass. This phosphorylation cycle is catalyzed by a bifunctional enzyme, IDH kinase/phosphatase, which is encoded by aceK. We have employed random mutagenesis to isolate novel alleles of aceK. These alleles were detected by the loss of ability to complement an aceK null mutation. The products of one class of these alleles retain IDH kinase activity but have suffered reductions in IDH phosphatase activity by factors of 200 to 400. Selective loss of the phosphatase activity also appears to have occurred in vivo, since cells expressing these alleles exhibit phenotypes which are reminiscent of strains lacking IDH; these strains are auxotrophic for glutamate. Assays of cell-free extracts confirmed that this phenotype resulted from nearly quantitative phosphorylation of IDH. The availability of these novel alleles of aceK allowed us to assess the significance of the precise control which is a characteristic of the IDH phosphorylation cycle in vivo. The fractional phosphorylation of IDH was varied by controlled expression of one of the mutant alleles, aceK3, in a wild-type strain. Reduction of IDH activity to 50% of the wild-type level did not adversely affect growth on acetate. However, further reductions inhibited growth, and growth arrest occurred when the IDH activity fell to 15% of the wild-type level. Thus, although wild-type cells maintain a precise effective IDH activity during growth on acetate, this precision is not critical. SN - 0021-9193 UR - https://www.unboundmedicine.com/medline/citation/1847910/Isocitrate_dehydrogenase_kinase/phosphatase:_aceK_alleles_that_express_kinase_but_not_phosphatase_activity_ L2 - https://journals.asm.org/doi/10.1128/jb.173.5.1801-1806.1991?url_ver=Z39.88-2003&rfr_id=ori:rid:crossref.org&rfr_dat=cr_pub=pubmed DB - PRIME DP - Unbound Medicine ER -