Determination of reaction volumes and polymer distribution characteristics of tobacco mosaic virus coat protein.
A method that allows the quantitative determination of reaction volumes from sedimentation velocity experiments in an analytical ultracentrifuge is presented. Combined with a second method for detecting pressure-induced depolymerization, general characteristics of polymer distributions may be probed. We show that it is possible to determine if a sample is in an equilibrium or metastable state of subunit association. Our approach to probe macromolecular aggregation systems by small pressure perturbations is not restricted to the use of centrifuges. This method has been applied to characterize certain aspects of the polymerization of tobacco mosaic virus coat protein (TMVP). There are at least two helical polymer conformations in RNA-free coat protein rods. The smaller, helix I, polymers are limited to sizes below about 70 subunits (four to five helical turns) and undergo some kind of cooperative conformational change before further subunits may be added indefinitely. In contrast to helix I, the larger helix II polymers occur as broader and skewed size distributions. Under moderately strong polymerization conditions, the equilibrium state can contain both types of helical rods. The reaction volume for the addition of trimers is -220 ml/mol for both types of helical polymers. These results are compared with the results of previous thermodynamic analyses of TMVP polymerization.
D002213:Capsid / Q000032:analysis*; D036022:Capsid Proteins; D011108:Polymers / Q000032:analysis; D011312:Pressure; D011487:Protein Conformation; D014027:Tobacco Mosaic Virus / Q000032:analysis*; D014461:Ultracentrifugation; D014764:Viral Proteins / Q000032:analysis*
D036022:Capsid Proteins; C009496:H protein, Tobacco mosaic virus; D011108:Polymers; D014764:Viral Proteins