Role of cardiac troponin I carboxy terminal mobile domain and linker sequence in regulating cardiac contraction.

Inhibition of striated muscle contraction at resting Ca(2+) depends on the C-terminal half of troponin I (TnI) in thin filaments. Much focus has been on a short inhibitory peptide (Ip) sequence within TnI, but structural studies and identification of disease-associated mutations broadened emphasis to include a larger mobile domain (Md) sequence at the C-terminus of TnI. For Md to function effectively in muscle relaxation, tight mechanical coupling to troponin's core-and thus tropomyosin-is presumably needed. We generated recombinant, human cardiac troponins containing one of two TnI constructs: either an 8-amino acid linker between Md and the rest of troponin (cTnILink8), or an Md deletion (cTnI1-163). Motility assays revealed that Ca(2+)-sensitivity of reconstituted thin filament sliding was markedly increased with cTnILink8 (∼0.9 pCa unit leftward shift of speed-pCa relation compared to WT), and increased further when Md was missing entirely (∼1.4 pCa unit shift). Cardiac Tn's ability to turn off filament sliding at diastolic Ca(2+) was mostly (61%), but not completely eliminated with cTnI1-163. TnI's Md is required for full inhibition of unloaded filament sliding, although other portions of troponin-presumably including Ip-are also necessary. We also confirm that TnI's Md is not responsible for superactivation of actomyosin cycling by troponin.