Calcium-dependent movement of troponin I between troponin C and actin as revealed by spin-labeling EPR

We measured EPR spectra from a spin label on the Cys133 residue of troponin I (TnI) to identity Ca2+-induced structural states, based on sensitivity of spin-label mobility to flexibility and tertiary contact of polypeptide. Spectrum from Tn Complexes in the -Ca2+ state showed that Cyst 33 was located at a flexible polypeptide segment (rotational correlation time tau 1.9 its) that was free from TnC. Spectra of both Tn complexes alone and those reconstituted into the thin filaments in the +Ca2+ state shocked that Cys133 existed on a stable segment (tau = 4.8 ns) held by TnC. Spectra of reconstituted thin filaments (-Ca2+ State) reveated that slow mobility (tau - 45 ns) was due to tertiary contact of Cys133 with actin, because the same slow mobility was found for TnI actin and TnI tropomyosin actin filaments lacking TnC, T or tropomyosin. We propose that the Cys133 region dissociates from TnC and attaches to the actin surface oil the thin filaments, causing muscle relaxation at low Ca2+ concentrations. (c) 2005 Elsevier Inc. All rights reserved.