title: Caldesmon freezes the structure of actin filaments during the actomyosin ATPase cycle.
abstract: Hybrid contractile apparatus was reconstituted in skeletal muscle ghost fibers by incorporation of skeletal muscle myosin subfragment 1 (S1), smooth muscle tropomyosin and caldesmon. The spatial orientation of FITC-phalloidin-labeled actin and IAEDANS-labeled S1 during sequential steps of the acto-S1 ATPase cycle was studied by measurement of polarized fluorescence in the absence or presence of nucleotides conditioning the binding affinity of both proteins. In the fibers devoid of caldesmon addition of nucleotides evoked unidirectional synchronous changes in the orientation of the fluorescent probes attached to F-actin or S1. The results support the suggestion on the multistep rotation of the cross-bridge (myosin head and actin monomers) during the ATPase cycle. The maximal cross-bridge rotation by 7 degrees relative to the fiber axis and the increase in its rigidity by 30% were observed at transition between A**.M**.ADP.Pi (weak binding) and A--.M--.ADP (strong binding) states. When caldesmon was present in the fibers (OFF-state of the thin filament) the unidirectional changes in the orientation of actin monomers and S1 were uncoupled. The tilting of the myosin head and of the actin monomer decreased by 29% and 90%, respectively. It is suggested that in the "closed" position caldesmon "freezes" the actin filament structure and induces the transition of the intermediate state of actomyosin towards the weak-binding states, thereby inhibiting the ATPase activity of the actomyosin.
authors.affiliation: Laboratory of Mechanisms of Cell Motility, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Avenue, St. Petersburg 194064, Russia.
authors.forename: Yurii S
authors.initials: YS
authors.lastname: Borovikov
authors.forename: Natalia
authors.initials: N
authors.lastname: Kulikova
authors.forename: Olga E
authors.initials: OE
authors.lastname: Pronina
authors.forename: Svetlana S
authors.initials: SS
authors.lastname: Khaimina
authors.forename: Antoni
authors.initials: A
authors.lastname: Wrzosek
authors.forename: Renata
authors.initials: R
authors.lastname: Dabrowska
mesh_terms: D000199:Actins / Q000737:chemistry*; D000205:Actomyosin / Q000737:chemistry*; D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000737:chemistry* / Q000502:physiology; D004372:Ducks; D018482:Muscle, Skeletal / Q000378:metabolism; D009130:Muscle, Smooth / Q000378:metabolism; D015879:Myosin Subfragments / Q000737:chemistry*; D009218:Myosins / Q000737:chemistry*; D010590:Phalloidine / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
chemical_list: D000199:Actins; D002148:Calmodulin-Binding Proteins; D015879:Myosin Subfragments; D010590:Phalloidine; D000205:Actomyosin; D009218:Myosins