A fluorescent probe study of Ca2+ binding to the Ca2+-specific sites of cardiac troponin and troponin C

J. D. Johnson, J. H. Collins, S. P. Robertson, J. D. Potter

Research output: Contribution to journalArticle

110 Scopus citations

Abstract

Cardiac troponin C (C-TnC) was labeled with the sulfhydryl-specific fluorescent probe molecule 2-(4'-iodoacetamidoanilino)naphthalene-6-sulfonic acid at cysteine 35 and 84 to produce C-TnC(IA). This modified protein binds Ca2+, undergoes Ca2+-induced increases in α helix, and forms a complex with other troponin subunits as does unlabeled C-TnC. C-TnC(IA) undergoes a small fluorescence decrease with Ca2+ or Mg2+ binding to the two high affinity Ca2+, Mg2+ sites of C-TnC and a large biphasic ~ 2.1-fold fluorescence increase with Ca2+ binding to two lower affinity Ca2+-specific sites with K(Ca) of ~ 4.5 x 10:5 M-1 and ~ 5 x 102 M-1. C-TnC(IA) was formed in a complex with troponin I (TnI) and troponin T to form C-Tn(IA). This fluorescent reconstituted whole troponin undergoes a 25% decrease with Ca2+ binding to a Ca2+ -specific site of K(Ca) ~ 3 x 106 M-1. C-TnC, therefore, contains a single Ca2+-specific site of approximate equal affinity as the two Ca2+-specific regulatory sites of skeletal TnC. This Ca2+-specific site in C-TnC (like its two corresponding sites in S-TnC) undergoes an approximate 10-fold increase in affinity in whole troponin or when TnC is complexed with TnI. Since the two Ca2+-specific sites in skeletal troponin have been shown to be the regulatory sites of skeletal muscle contraction we suggest that this single Ca2+-specific site, of equal affinity, in C-TnC is the regulatory site of cardiac muscle contraction.

Original languageEnglish (US)
Pages (from-to)9635-9640
Number of pages6
JournalJournal of Biological Chemistry
Volume255
Issue number20
StatePublished - Dec 1 1980

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Fingerprint Dive into the research topics of 'A fluorescent probe study of Ca<sup>2+</sup> binding to the Ca<sup>2+</sup>-specific sites of cardiac troponin and troponin C'. Together they form a unique fingerprint.

  • Cite this

    Johnson, J. D., Collins, J. H., Robertson, S. P., & Potter, J. D. (1980). A fluorescent probe study of Ca2+ binding to the Ca2+-specific sites of cardiac troponin and troponin C. Journal of Biological Chemistry, 255(20), 9635-9640.