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P Vibert

Publications and source records attributed to P Vibert.

27 records · Page 2Linked to original sources
PubMed1985

Structural changes that occur in scallop myosin filaments upon activation.

Myosin filaments isolated from scallop striated muscle have been activated by calcium-containing solutions, and their structure has been examined by electron microscopy after negative staining. The orderly helical arrangement of myosin projections characteristic of the relaxed state is largely lost upon activation. The oblique striping that arises from alignment of elongated projections along the long-pitched helical tracks is greatly weakened, although a 145 A axial periodicity is sometimes partially retained. The edges of the filaments become rough, and the myosin heads move outwards as their helical arrangement becomes disordered. Crossbridges at various angles appear to link thick and thin filaments after activation. The transition from order to disorder is reversible and occurs over a narrow range of free calcium concentration near pCa 5.7. Removal of nucleotide, as well as dissociation of regulatory light chains, also disrupts the ordered helical arrangement of projections. We suggest that the relaxed arrangement of the projections is probably maintained by intermolecular interactions between myosin molecules, which depend on the regulatory light chains. Calcium binding changes the interactions between light chains and the rest of the head, activating the myosin molecule. Intermolecular contacts between molecules may thus be altered and may propagate activation cooperatively throughout the thick filament.

Adenosine Triphosphate↗
PubMed1983

Electron microscopy of scallop myosin. Location of regulatory light chains.

The heads of the Ca2+-sensitive myosin molecules from scallop muscle, contrasted for electron microscopy by rotary shadowing, display two appearances depending on the presence or absence of the regulatory light chains. The heads of intact myosin appear "pear-shaped" as described for vertebrate myosin (Elliott & Offer, 1978): they are widest at the end remote from the tail and taper to a narrower neck near their junction with the tail. In contrast, myosin heads that lack the regulatory light chains appear more globular. The neck region is no longer visible: the rounded heads appear directly attached to the tail or there is an apparent gap between the head and the tail. Two preparations of myosin subfragment-1 that differ in light chain content show a similar difference in appearance. Fab fragments of antibodies specific for the light chains bind to the myosin heads and can also be visualized in the electron microscope using rotary shadowing. Both Fab fragments specific for the regulatory light chains and Fab fragments specific for the essential light chains bind preferentially to intact scallop myosin in the narrow region of the myosin head near its junction with the tail.

Animals↗
PubMed1983

Structure of myosin/paramyosin filaments from a molluscan smooth muscle.

Small-angle X-ray diffraction patterns of chemically skinned pedal retractor muscles from Mytilus (PRM) in the relaxed state show a set of diffuse off-meridional reflections that arise from a helical array of myosin crossbridges with 8/3 screw symmetry. Experiments involving extraction of myosin as well as analysis of the rigor pattern have been used to confirm the origin of these reflections. The relatively high myosin/paramyosin molar ratio (1.3 to 1.6) in PRM compared to other molluscan smooth muscles may account for the observation of the relatively stronger diffraction from the myosin array. Thick filaments isolated from PRM and contrasted by negative staining for electron microscopy appear to be very long (up to 17 micron), and to have a rather small diameter (about 40 nm at the center); they taper gradually toward the ends. These filaments show a clear transverse band pattern repeating at 14.4 nm and elongated projections (crossbridges) at the surface except in the central bare zone. Optical diffraction patterns show reflections from crossbridges consistent with the X-ray patterns of the relaxed whole muscle. Filaments unidirectionally shadowed with platinum show diagonal striations running at an angle of about 17 degrees to the filament axis, revealing that the crossbridges are arrayed in a right-handed helix. The paramyosin core is clearly seen upon extraction of myosin. Observations on both negatively stained and sectioned material are consistent with the results of Elliott (1979) and Bennett & Elliott (1981) suggesting a layered structure of the core. Cores stripped of myosin, however, appear to undergo some distortion indicating that the three-dimensional structure is not yet completely solved. The assembly of these thick filaments presents intriguing structural problems since the myosin surface lattice does not appear to have the same symmetry as the underlying paramyosin core.

Animals↗
PubMed1983

Electron microscopy and image analysis of myosin filaments from scallop striated muscle.

Thick filaments have been isolated from the striated adductor muscle of the scallop and examined by electron microscopy after negative staining. Many filaments appear intact, and reveal a centrally located bare-zone and a well-defined helical surface array of myosin crossbridges characterized by a 145 A axial period and prominent helical tracks of pitch 480 A. Heavy-metal shadowing shows that these helices are right-handed. A small perturbation of alternate crossbridge levels produces an axial period of 290 A, which is most prominent in a region on either side of the bare-zone. Image analysis reveals that the crossbridge array has 7-fold rotational symmetry, one of the possibilities suggested by earlier X-ray diffraction studies of native filaments in scallop muscle. A low-resolution three-dimensional reconstruction shows elongated surface projections ("crossbridges") that probably represent unresolved pairs of myosin heads. They run almost parallel to the filament surface, but are slewed slightly from the axis so that they lie along the right-handed helical tracks of pitch 480 A. The connection to the filament backbone probably occurs at the end of the crossbridges nearer the bare-zone; thus, their sense of tilt appears to be opposite to that of rigor attachment to actin. The 290 A period arises from a different distribution of crossbridge density at alternate levels; in addition, there are weak connections between the top of one crossbridge and the bottom of the next, 145 A away. The prominence of the 290 A period near the bare-zone suggests that anti-parallel molecular interactions are mainly responsible for this perturbation.

Animals↗
PubMedDate not supplied

A new myosin fragment: visualization of the regulatory domain.

Many of the functional domains of the myosin molecule have been defined by the use of proteolytic enzymes. Major fragments that retain enzymatic or assembly properties have been prepared by cleavage in the rod to form heavy meromyosin (HMM) and light meromyosin (LMM) or at the head-rod junction to form S-1 and rod. Limited tryptic digestion of vertebrate skeletal myosin S-1 indicates that the head contains three major regions: an amino-terminal nucleotide binding domain of molecular weight (MW) 25,000, a central domain of MW 50,000 and a carboxyl domain MW 20,000; the latter two are both able to bind to actin. Tryptic digestion of scallop S-1 has also been used to isolate a head fragment MW 14,000 associated with both types of scallop light chains. Here we report that myosin from vertebrate (chicken and rabbit skeletal) and molluscan (scallop adductor) striated muscles is cleaved in an unusual way with an enzyme from Pseudomonas aeruginosa. This bacterial protease (designated Ps-1) does not cleave myosin at the head-rod junction or in the rod; instead, Ps-1 splits the myosin heavy chain within the head, yielding a complete rod joined to the 20,000-MW head domains. The scallop regulatory and essential light chains remain associated with this fragment. We examined this new fragment by electron microscopy; the rods bear two 'nubs' about 100 A long, which appear to correspond morphologically to the neck region of the myosin molecule.

Animals↗