Investigation of the mechanism of calcium binding to calmodulin. Use of an isofunctional mutant with a tryptophan introduced by site-directed mutagenesis

M. C. Kilhoffer, D. M. Roberts, A. O. Adibi, D. M. Watterson, J. Haiech

Research output: Contribution to journalArticle

29 Scopus citations

Abstract

A mutant calmodulin, in which phenylalanine 99 of calcium binding site III was changed to a tryptophan by using cassette-based, site-directed mutagenesis, has been used to analyze the mechanism of calcium binding. The combined study of direct calcium binding, modification of tryptophan fluorescence properties upon calcium binding, and terbium titration allows some discrimination among proposed mechanisms of cation binding to calmodulin. Calmodulin appears to have six cation binding sites, four of which are selective for calcium that seem to be coupled. Under a given set of conditions, these calcium-selective sites are not identical. In addition to providing insight into the mechanisms of calcium modulation of calmodulin, these studies demonstrate the feasibility of using isofunctional, tryptophan-containing mutants of proteins to gain insight into protein-ligand interaction.

Original languageEnglish (US)
Pages (from-to)17023-17029
Number of pages7
JournalJournal of Biological Chemistry
Volume263
Issue number32
StatePublished - 1988

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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