The trout heart is 10-fold more sensitive to Ca 2+ than the mammalian heart. This difference is due, in part, to cardiac troponin C (cTnC) from trout having a greater Ca 2+ affinity than human cTnC. To determine what other proteins are involved, we cloned cardiac troponin I (cTnI) from the trout heart and determined how it alters the Ca 2+ affinity of a cTn complex containing all mammalian components (mammalian cTn). Ca 2+ activation of the complex was characterized using a human cTnC mutant

more »

... t contains anilinonapthalenesulfote iodoacetamide attached to Cys53. When the cTn complex containing labeled human cTnC was titrated with Ca 2+ , its fluorescence changed, reaching an asymptote upon saturation. Our results reveal that trout cTnI lacks the N-terminal extension found in cTnI from all other vertebrate groups. This protein domain contains two targets (Ser23 and Ser24) for protein kinase A (PKA) and protein kinase C. When these are phosphorylated, the rate of cardiomyocyte relaxation increases. When rat cTnI in the mammalian cTn complex was replaced with trout cTnI, the Ca 2+ affinity was increased ~1.8-fold. This suggests that trout cTnI contributes to the high Ca 2+ sensitivity of the trout heart. Treatment of the two cTn complexes with PKA decreased the Ca 2+ affinity of both complexes. However, the change for the complex containing rat cTnI was 2.2-fold that of the complex containing trout cTnI. This suggests that the phosphorylation of trout cTnI does not play as significant a role in regulating cTn function in trout.