Actin-attached and detached crossbridges in myofibrils: segregation into two populations according to their sensitivity to proteolytic digestion of myosin heavy chain.
Tryptic digestion of myofibrils was used to assess the interaction of crossbridges with thin filaments in the presence of ATP analogues. The relative amounts of 200 kDa fragment produced by trypsin from myosin heavy chain when the crossbridge is attached to actin, and of 160 kDa fragment produced when the crossbridge is detached from actin, served as a measure of crossbridge-actin interaction. In rigor only the 200 kDa fragment was produced suggesting that a great majority of the crossbridges were strongly attached to actin; in the presence of MgPPi at 0 degrees C only the 160 kDa fragment was finally produced suggesting that eventually all crossbridges detached from actin. In the presence of MgPPi or MgAMPPNP at 25 degrees C both 200 and 160 kDa fragments were present for several minutes after myosin heavy chain had been completely digested, suggesting that two populations of crossbridges (attached and detached) co-existed at the same time within the myofibril. It is concluded that the addition of ATP analogues to muscle does not simply affect the chemical equilibrium of binding of myosin heads to actin but that it causes rapid dissociation of one crossbridge population without significant effect on binding to actin of the remaining crossbridge population.
D000199:Actins / Q000032:analysis*; D000266:Adenylyl Imidodiphosphate / Q000494:pharmacology; D000818:Animals; D011756:Diphosphates / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D008024:Ligands; D008274:Magnesium / Q000494:pharmacology; D017616:Magnesium Compounds; D008970:Molecular Weight; D009210:Myofibrils / Q000032:analysis*; D009218:Myosins / Q000032:analysis*; D010446:Peptide Fragments / Q000032:analysis; D011817:Rabbits; D013696:Temperature; D014357:Trypsin / Q000494:pharmacology
D000199:Actins; D011756:Diphosphates; D008024:Ligands; D017616:Magnesium Compounds; D010446:Peptide Fragments; C033025:actin subfragments; D000266:Adenylyl Imidodiphosphate; C034732:magnesium pyrophosphate; D014357:Trypsin; D009218:Myosins; D008274:Magnesium