Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1973 Nov;116(2):790–796. doi: 10.1128/jb.116.2.790-796.1973

Studies on the Enzymes Involved in the Biosynthesis of Cyclo-Tris (N-2,3-Dihydroxybenzoyl-l-Seryl) in Escherichia coli: Kinetic Properties of the l-Serine–Activating Enzyme

Graeme F Bryce 1
PMCID: PMC285447  PMID: 4126827

Abstract

Some kinetic properties of the formation of a thioester-bound l-seryl-enzyme intermediate are described. The rate constant of formation is 3.12 × 102 M−1 s−1 and the rate constant for spontaneous breakdown is 2.47 × 10−3 s−1. These constants yield a value of log K = 5.10 for the overall equilibrium constant which agrees favorably with a value of 5.20 calculated from equilibrium binding data. E1·serine formation requires a thiol group which is extremely reactive to N-ethylmaleimide; the second-order rate constant for enzyme inactivation by this reagent is 77.1 M−1 s−1 at pH 6.6 and 0 C. Excess l-serine does not protect the enzyme against inactivation. In addition to an adenosine triphosphate-[32P]-inorganic pyrophosphate exchange, the enzyme also catalyzes an l-serine-dependent adenosine triphosphate-[3H]adenosine monophosphate exchange in accordance with the scheme proposed for the activation of serine.

Full text

PDF
790

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson B. M., Vasini E. C. Nonpolar effects in reactions of the sulfhydryl group of papain. Biochemistry. 1970 Aug 18;9(17):3348–3352. doi: 10.1021/bi00819a009. [DOI] [PubMed] [Google Scholar]
  2. Brot N., Goodwin J., Fales H. In vivo and in vitro formation of 2,3-dihydroxybenzoylserine by Escherichia coli K12. Biochem Biophys Res Commun. 1966 Nov 22;25(4):454–461. doi: 10.1016/0006-291x(66)90227-0. [DOI] [PubMed] [Google Scholar]
  3. Brot N., Goodwin J. Regulation of 2,3-dihydroxybenzoylserine synthetase by iron. J Biol Chem. 1968 Feb 10;243(3):510–513. [PubMed] [Google Scholar]
  4. Bryce G. F., Brot N. Iron transport in Escherichia coli and its relation to the repression of 2,3-dihydroxy-N-benzoyl-L-serine synthetase. Arch Biochem Biophys. 1971 Feb;142(2):399–406. doi: 10.1016/0003-9861(71)90503-0. [DOI] [PubMed] [Google Scholar]
  5. Bryce G. F., Brot N. Studies on the enzymatic synthesis of the cyclic trimer of 2,3-dihydroxy-N-benzoyl-L-serine in Escherichia coli. Biochemistry. 1972 Apr 25;11(9):1708–1715. doi: 10.1021/bi00759a028. [DOI] [PubMed] [Google Scholar]
  6. Bryce G. F., Weller R., Brot N. Studies on the enzymatic synthesis of 2,3-dihydroxy-N-benzoyl-L-serine in Escherichia coli. Biochem Biophys Res Commun. 1971 Mar 5;42(5):871–879. doi: 10.1016/0006-291x(71)90511-0. [DOI] [PubMed] [Google Scholar]
  7. Colman R. F., Chu R. The role of sulfhydryl groups in the catalytic function of isocitrate dehydrogenase. II. Effect of N-ethylmaleimide on kinetic properties. J Biol Chem. 1970 Feb 10;245(3):601–607. [PubMed] [Google Scholar]
  8. Gevers W., Kleinkauf H., Lipmann F. Peptidyl transfers in gramicidin S bisoynthesis from enzyme-bound thioester intermediates. Proc Natl Acad Sci U S A. 1969 Aug;63(4):1335–1342. doi: 10.1073/pnas.63.4.1335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Gevers W., Kleinkauf H., Lipmann F. The activation of amino acids for biosynthesis of gramicidin S. Proc Natl Acad Sci U S A. 1968 May;60(1):269–276. doi: 10.1073/pnas.60.1.269. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Katze J. R., Konigsberg W. Purification and properties of seryl transfer ribonucleic acid synthetase from Escherichia coli. J Biol Chem. 1970 Mar 10;245(5):923–930. [PubMed] [Google Scholar]
  11. Krishna G., Weiss B., Brodie B. B. A simple, sensitive method for the assay of adenyl cyclase. J Pharmacol Exp Ther. 1968 Oct;163(2):379–385. [PubMed] [Google Scholar]
  12. O'Brien I. G., Cox G. B., Gibson F. Biologically active compounds containing 2,3-dihydroxybenzoic acid and serine formed by Escherichia coli. Biochim Biophys Acta. 1970 Mar 24;201(3):453–460. doi: 10.1016/0304-4165(70)90165-0. [DOI] [PubMed] [Google Scholar]
  13. Roskoski R., Jr, Gevers W., Kleinkauf H., Lipmann F. Tyrocidine biosynthesis by three complementary fractions from Bacillus brevis (ATCC 8185). Biochemistry. 1970 Dec 8;9(25):4839–4845. doi: 10.1021/bi00827a002. [DOI] [PubMed] [Google Scholar]
  14. Roskoski R., Jr, Kleinkauf H., Gevers W., Lipmann F. Isolation of enzyme-bound peptide intermediates in tyrocidine biosynthesis. Biochemistry. 1970 Dec 8;9(25):4846–4851. doi: 10.1021/bi00827a003. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES