What does TnCDANZ fluorescence reveal about the thin filament state?
Fluorescence of skinned psoas fibres reconstituted with the troponin C subunit labelled with the fluorescent probe dansylaziridine (TnCDANZ) increases upon activation with Ca2+. This fluorescence enhancement is due to Ca2+ binding to the Ca(2+)-specific binding sites of TnCDANZ and attachment of cross-bridges to the actin filament. We found that approximately 20% of the enhanced fluorescence signal derived from Ca2+ binding to TnCDANZ and 80% from cross-bridge attachment during maximal activation. Furthermore we studied the effects of different cross-bridge states on TnCDANZ fluorescence. "Weakly" bound, non-force-generating cross-bridge states (pCa 8, low ionic strength) and rigor cross-bridges revealed similar effects on the TnCDANZ fluorescence. "Strongly" attached, force-generating states, however, increased fluorescence to the greatest extent. These results suggests a complex system of reciprocal couplings between TnC and different attached cross-bridge states. Cooling or increase of inorganic phosphate decreased isometric force but hardly decreased fluorescence, suggesting the accumulation of attached cross-bridge states with low tension output.
Morano
I
I
Department of Physiology II, University of Heidelberg, Federal Republic of Germany.
Rüegg
J C
JC
D008841:Actin Cytoskeleton / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D000818:Animals; D001388:Aziridines; D002118:Calcium / Q000378:metabolism; D003619:Dansyl Compounds; D005456:Fluorescent Dyes; D007537:Isometric Contraction / Q000502:physiology; D009119:Muscle Contraction / Q000502:physiology; D009132:Muscles / Q000378:metabolism* / Q000502:physiology / Q000648:ultrastructure; D011817:Rabbits; D014336:Troponin / Q000378:metabolism*; D019209:Troponin C
D001388:Aziridines; D003619:Dansyl Compounds; D005456:Fluorescent Dyes; D014336:Troponin; D019209:Troponin C; C017173:dansylaziridine; D002118:Calcium