title: Cryoenzymic studies on actomyosin ATPase: kinetic evidence for communication between the actin and ATP sites on myosin.
abstract: The post-ATP binding steps of myosin subfragment 1 (S1) and actomyosin subfragment 1 (actoS1) ATPases were studied at -15 degrees C with 40% ethylene glycol as antifreeze. The cleavage and release of Pi steps were studied by the rapid-flow quench method and the interaction of actin with S1 plus ATP by light scattering in a stopped-flow apparatus. At -15 degrees C, the interaction of actin with S1 remains tight, and the Km for the activation of S1 ATPase is very small (0.3 microM). The chemical data were interpreted by E + ATP----E*.ATP----E**.ADP.Pi----E*.ADP----products, where E is S1 or actoS1. In Pi burst experiments with S1, there was a large Pi burst of free Pi, but E**.ADP.Pi could not be detected. Here the predominant complex in the seconds time range is E*.ATP and in the steady-state E*.ADP. With actoS1, there was a small Pi burst of E**.ADP.Pi, evidence that the cleavage steps for S1 and actoS1 are different. From the stopped-flow experiments, the dissociation of actoS1 by ATP was complete, even at actin concentrations 60X its Km. Further, no interaction of actin with the key intermediate M*.ATP could be detected. Therefore, at -15 degrees C, actoS1 ATPase occurs by a dissociative pathway; in particular, the cleavage step appears to occur in the absence of actin.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: INSERM U128, CNRS, BP 5051, Montpellier, France.
authors.forename: C
authors.initials: C
authors.lastname: Tesi
authors.forename: K
authors.initials: K
authors.lastname: Kitagishi
authors.forename: F
authors.initials: F
authors.lastname: Travers
authors.forename: T
authors.initials: T
authors.lastname: Barman
mesh_terms: D000199:Actins / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001665:Binding Sites; D019855:Ethylene Glycol; D005026:Ethylene Glycols; D005615:Freezing; D007700:Kinetics; D015879:Myosin Subfragments / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D011485:Protein Binding; D013816:Thermodynamics
chemical_list: D000199:Actins; D005026:Ethylene Glycols; D015879:Myosin Subfragments; D000255:Adenosine Triphosphate; D009218:Myosins; D019855:Ethylene Glycol