Calcium-induced flexibility changes in the troponin C-troponin I complex

The contraction of vertebrate striated muscle is modulated by Ca2+ binding to the regulatory protein troponin C (TnC). Ca2+ binding causes conformatio nal changes in TnC which alter its interaction with the inhibitory protein troponin I (TnI), initiating the regulatory process. We have used the frequ ency domain method of fluorescence resonance energy transfer (FRET) to meas ure distances and distance distributions between specific sites in the TnC- TnI complex in the presence and absence of Ca2+ or Mg2+. Using sequences ba sed on rabbit skeletal muscle proteins, we prepared functional, binary comp lexes of wild-type TnC and a TnI mutant which contains no Cys residues and a single Trp residue at position 106 within the TnI inhibitory region. We u sed TnI Trp-106 as the FRET donor, and we introduced energy acceptor groups into TnC by labeling at Met-25 with dansyl aziridine or at Cys-98 with N-( iodoacetyl)-N'-(1-sulfo-5-naphthyl)ethylenediamine. Our distance distributi on measurements indicate that the TnC-TnI complex is relatively rigid in th e absence of Ca2+, but becomes much more flexible when Ca2+ binds to regula tory sites in TnC. This increased flexibility may be propagated to the whol e thin filament, helping to release the inhibition of actomyosin ATPase act ivity and allowing the muscle to contract. This is the first report of dist ance distributions between TnC and TnI in their binary complex. (C) 2000 El sevier Science B.V. All rights reserved.