Stopped-flow, classical, and dynamic light scattering analysis of matrix protein binding to nucleocapsids of vesicular stomatitis virus.
During the process of assembly of enveloped viruses, binding of the nucleoprotein core of the virus (nucleocapsid) to the host membrane is mediated by the viral matrix (M) protein. Light scattering properties of vesicular stomatitis virus (VSV) nucleocapsids and nucleocapsid-M protein (NCM) complexes assembled in vivo were determined following solubilizaton of the virion envelope with detergents at varying ionic strength to vary the extent of M protein binding. Three factors were found to contribute to the light scattering properties of VSV nucleocapsids: their conformation, extent of self-association, and amount of bound M protein. All three were affected by changes in ionic strength but could be distinguished by several parameters. Conformational changes in nucleocapsids and NCM complexes occurred rapidly (millisecond time scale) upon changing salt concentration and were reflected in changes in the angular dependence of light scattering intensity (i.e., changes in radius of gyration, RG). Changes in extent of self-association occurred relatively slowly (seconds to minutes time scale) and could be distinguished by the concentration dependence of the apparent molecular mass and diffusion coefficient of the NCM complex. Changes in M protein binding occurred on an intermediate time scale (t1/2 approximately one s) and reflected changes in both molecular mass and RG. The data presented here provide criteria for assessing binding of M protein to nucleocapsids under conditions of minimal perturbation of the NCM complex assembled in vivo and at low protein concentrations so that self-association of the NCM complex was minimal and reversible.
Lyles
D S
DS
Department of Microbiology and Immunology, Bowman Gray School of Medicine of Wake Forest University, Winston-Salem, North Carolina 27157-11064, USA.
McKenzie
M O
MO
Hantgan
R R
RR
D000818:Animals; D002213:Capsid / Q000737:chemistry* / Q000378:metabolism; D002460:Cell Line; D006224:Cricetinae; D007700:Kinetics; D008027:Light; D046911:Macromolecular Substances; D008970:Molecular Weight; D011485:Protein Binding; D011487:Protein Conformation; D012542:Scattering, Radiation; D012965:Sodium Chloride; D014721:Vesicular stomatitis Indiana virus / Q000737:chemistry* / Q000254:growth & development / Q000378:metabolism; D014763:Viral Matrix Proteins / Q000737:chemistry* / Q000378:metabolism
D046911:Macromolecular Substances; D014763:Viral Matrix Proteins; D012965:Sodium Chloride