title: Characterization of myosin V binding to brain vesicles.
abstract: Myosin II and V are important for the generation and segregation of subcellular compartments. We observed that vesicular myosin II and V were associated with the protein scaffolding of a common subset of vesicles by density sedimentation, electron microscopy, and immunofluorescence. Solubilization of either myosin II or V was caused by polyphosphates with the following efficacy at 10 mM: for myosin II ATP-Mg(2+) = ATP = AMP-PNP (5'-adenylyl imidodiphosphate) > pyrophosphate = tripolyphosphate > tetrapolyphosphate = ADP > cAMP = Mg(2+); and for myosin V pyrophosphate = tripolyphosphate > ATP-Mg(2+) = ATP = AMP-PNP > ADP = tetrapolyphosphate > cAMP = Mg(2+). Consequently, we suggest solubilization was not an effect of phosphorylation, hydrolysis, or disassociation of myosin from actin filaments. Scatchard analysis of myosin V binding to stripped dense vesicles showed saturable binding with a K(m) of 10 nM. Analysis of native vesicles indicates that these sites are fully occupied. Together, these data show there are over 100 myosin Vs/vesicle (100-nm radius). We propose that polyphosphate anions bind to myosin II and V and induce a conformational change that disrupts binding to a receptor.
authors.affiliation: Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: K E
authors.initials: KE
authors.lastname: Miller
authors.forename: M P
authors.initials: MP
authors.lastname: Sheetz
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000196:embryology / Q000378:metabolism*; D002642:Chick Embryo; D000242:Cyclic AMP / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism; D008854:Microscopy, Electron; D009218:Myosins / Q000378:metabolism* / Q000648:ultrastructure; D010710:Phosphates / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms / Q000378:metabolism* / Q000648:ultrastructure; D012965:Sodium Chloride
chemical_list: D010710:Phosphates; D020033:Protein Isoforms; D012965:Sodium Chloride; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D009218:Myosins