title: Cryo-electron microscopy of S1-decorated actin filaments.
abstract: We have applied techniques for cryo-electron microscopy, combined with image processing, to both S1-decorated native thin filaments and S1-decorated actin filaments. In our reconstruction the actin subunit has a prolate ellipsoid shape and is composed of two domains. The long axis of the monomer lies roughly perpendicular to the filament axis. The myosin head (S1) approaches the actin filament tangentially, the major interaction being with the outermost domain of actin. To distinguish the position of tropomyosin unambiguously in our map, we compared the maps from decorated thin filaments with those from decorated actin filaments. Our difference map clearly shows a peak corresponding to the position of tropomyosin; tropomyosin is bound to the inner domain of actin just in front of the myosin binding site at a radius of about 40 A. As a first step toward looking at the actomyosin structure in a state other than rigor, we examined S1 crosslinked to actin filaments by the zero-length crosslinker EDC in the presence of ATP and after pPDM bridging of the reactive thiols of S1. S1 molecules of the cross-linked complexes in the presence of ATP and after pPDM treatment appear dramatically different from those in rigor. The S1s appear more disordered and no longer assume the characteristic rigor 45 degrees angle with the actin filaments.
authors.affiliation: Department of Molecular Biology, Vanderbilt University, Nashville, TN 37235.
authors.forename: P F
authors.initials: PF
authors.lastname: Flicker
authors.forename: R A
authors.initials: RA
authors.lastname: Milligan
authors.forename: D
authors.initials: D
authors.lastname: Applegate
mesh_terms: D000199:Actins / Q000378:metabolism / Q000648:ultrastructure*; D000818:Animals; D001665:Binding Sites; D005615:Freezing; D008854:Microscopy, Electron / Q000379:methods; D008961:Models, Structural; D009132:Muscles / Q000648:ultrastructure*; D015879:Myosin Subfragments / Q000378:metabolism / Q000648:ultrastructure*; D011487:Protein Conformation; D014335:Tropomyosin / Q000648:ultrastructure
chemical_list: D000199:Actins; D015879:Myosin Subfragments; D014335:Tropomyosin