Sequence mutations in teleost cardiac troponin C that are permissive of high Ca2+ affinity of site II.
Cardiac myofibrils isolated from trout heart have been demonstrated to have a higher sensitivity for Ca(2+) than mammalian cardiac myofibrils. Using cardiac troponin C (cTnC) cloned from trout and mammalian hearts, we have previously demonstrated that this comparatively high Ca(2+) sensitivity is due, in part, to trout cTnC (ScTnC) having twice the Ca(2+) affinity of mammalian cTnC (McTnC) over a broad range of temperatures. The amino acid sequence of ScTnC is 92% identical to McTnC. To determine the residues responsible for the high Ca(2+) affinity, the function of a number of ScTnC and McTnC mutants was characterized by monitoring an intrinsic fluorescent reporter that monitors Ca(2+) binding to site II (F27W). The removal of the COOH terminus (amino acids 90-161) from ScTnC and McTnC maintained the difference in Ca(2+) affinity between the truncated cTnC isoforms (ScNTnC and McNTnC). The replacement of Gln(29) and Asp(30) in ScNTnC with the corresponding residues from McNTnC, Leu and Gly, respectively, reduced Ca(2+) affinity to that of McNTnC. These results demonstrate that Gln(29) and Asp(30) in ScTnC are required for the high Ca(2+) affinity of site II.
Gillis
Todd E
TE
Department of Biological Sciences, Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada.
Moyes
Chris D
CD
Tibbits
Glen F
GF
D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism*; D005399:Fishes / Q000235:genetics*; D006863:Hydrogen-Ion Concentration; D008322:Mammals / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation / Q000502:physiology*; D009206:Myocardium / Q000378:metabolism*; D017686:Oncorhynchus mykiss / Q000235:genetics; D010446:Peptide Fragments / Q000235:genetics; D013045:Species Specificity; D013696:Temperature; D019209:Troponin C / Q000235:genetics* / Q000378:metabolism*
D010446:Peptide Fragments; D019209:Troponin C; D002118:Calcium