Increasing mammalian cardiomyocyte contractility with residues identified in trout troponin C.
The Ca2+ sensitivity of force generation in trout cardiac myocytes is significantly greater than that from mammalian hearts. One mechanism that we have suggested to be responsible, at least in part, for this high Ca2+ sensitivity is the isoform of cardiac troponin C (cTnC) found in trout hearts (ScTnC), which has greater than twice the Ca2+ affinity of mammalian cTnC (McTnC). Here, through a series of mutations, the residues in ScTnC responsible for its high Ca2+ affinity have been identified as being Asn2, Ile28, Gln29, and Asp30. When these residues in McTnC were mutated to the trout-equivalent amino acid, the Ca2+ affinity of the molecule, determined by monitoring the fluorescence of a Trp inserted for a Phe at residue 27, is comparable to that of ScTnC. To determine how a McTnC mutant containing Asn2, Ile28, Gln29, and Asp30 (NIQD McTnC) affects the Ca2+ sensitivity of force generation, endogenous cTnC in single, chemically skinned rabbit cardiomyocytes was replaced with either wild-type McTnC or NIQD McTnC. Our results demonstrate that the cardiomyocytes containing NIQD McTnC were approximately twice as sensitive to Ca2+, illustrating that a McTnC mutant with similar Ca2+ affinity as ScTnC can be used to sensitize mammalian cardiac myocytes to Ca2+.
Gillis
Todd E
TE
Department of Bioengineering, University of Washington, Seattle, Washington, USA.
Liang
Bo
B
Chung
Franca
F
Tibbits
Glen F
GF
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009200:Myocardial Contraction / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000502:physiology*; D011485:Protein Binding; D011817:Rabbits; D016415:Sequence Alignment; D013997:Time Factors; D017186:Titrimetry; D019209:Troponin C / Q000737:chemistry* / Q000378:metabolism*; D014337:Trout
D000596:Amino Acids; D019209:Troponin C; D002118:Calcium