Calcium-bindings of wild type and mutant troponin Cs of Caenorhabditis elegans

Biochim Biophys Acta. 2001 Aug 13;1548(2):220-8. doi: 10.1016/s0167-4838(01)00234-5.

Abstract

Apparent Ca(2+)-binding constant (K(app)) of Caenorhabditis elegans troponin C (CeTnC) was determined by a fluorescence titration method. The K(app) of the N-domain Ca(2+)-binding site of CeTnC was 7.9+/-1.6 x 10(5) M(-1) and that of the C-domain site was 1.2+/-0.6 x 10(6) M(-1), respectively. Mg(2+)-dependence of the K(app) showed that both Ca(2+)-binding sites did not bind competitively Mg(2+). The Ca(2+) dissociation rate constant (k(off)) of CeTnC was determined by the fluorescence stopped-flow method. The k(off) of the N-domain Ca(2+)-binding site of CeTnC was 703+/-208 s(-1) and that of the C-domain site was 286+/-33 s(-1), respectively. From these values we could calculate the Ca(2+)-binding rate constant (k(on)) as to be 5.6+/-2.8 x 10(8) M(-1) s(-1) for the N-domain site and 3.4+/-2.1 x 10(8) M(-1) s(-1) for the C-domain site, respectively. These results mean that all Ca(2+)-binding sites of CeTnC are low affinity, fast dissociating and Ca(2+)-specific sites. Evolutional function of TnC between vertebrate and invertebrate and biological functions of wild type and mutant CeTnCs are discussed.

MeSH terms

  • Aminoquinolines
  • Animals
  • Binding Sites
  • Caenorhabditis elegans / metabolism*
  • Calcium / metabolism*
  • Evolution, Molecular
  • Fluorescent Dyes
  • Kinetics
  • Mutation
  • Spectrometry, Fluorescence
  • Titrimetry
  • Troponin C / genetics
  • Troponin C / metabolism*

Substances

  • Aminoquinolines
  • Fluorescent Dyes
  • Troponin C
  • Quin2
  • Calcium