Evidence for a small catalytic domain in the adenylate cyclase from Salmonella typhimurium.

T. K. Leib, J. A. Gerlt

Research output: Contribution to journalArticlepeer-review

Abstract

Deletions of large portions of the carboxyl-terminal end of the adenylate cyclase (ATP pyrophosphate lyase (cyclizing), EC 4.6.1.1) from Salmonella typhimurium do not significantly affect the enzymatic activity exhibited by the shortened polypeptide. The deletion mutations were generated by nuclease Bal31 digestion from the 3'-end of the cya gene fragment cloned by Wang et al. (Wang, J. Y.-J., Clegg, D. O., and Koshland, D.E. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 4684-4688); the shortened cya genes were inserted in pBR322 and used to transform a cya- strain of Escherichia coli. The original gene fragment encodes for an enzymatically active polypeptide having an apparent molecular weight of 77,000. Mutant polypeptides as small as 46,000 Da were found to retain significant enzymatic activity and to confer several cya+ phenotypes on the E. coli host. More extensive deletions resulting in polypeptides as small as 33,000 Da did not have assayable amounts of adenylate cyclase activity, but the biochemical properties of the transformed cya- host implicate the presence of low levels of enzymatic activity. These data suggest that the structure of the intact enzyme is composed of discrete functional domains. Such a structure for this adenylate cyclase should both facilitate investigations of the chemical mechanism of the reaction and allow structure-function relationships in this physiologically important enzyme to be investigated on a molecular level.

Original languageEnglish (US)
Pages (from-to)12982-12987
Number of pages6
JournalJournal of Biological Chemistry
Volume258
Issue number21
StatePublished - Nov 10 1983
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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