Biochemical evidence for the oligomeric (possibly trimeric) structure of the major inner capsid polypeptide (45K) of rotaviruses.

Abstract

The major structural polypeptide of rotaviruses is p45K (VP6), which forms the morphological subunit of the inner capsid. Such subunits show a trimeric structure when examined with the electron microscope. Treatment of single-capsid rotavirus particles with 1.5 M-CaCl2 removes p45K, resulting in the generation of smooth cores. Sucrose density gradient centrifugation analysis of the removed p45K revealed that it has a sedimentation coefficient close to 7.3S, compatible with an oligomeric (possibly trimeric) structure. Polyacrylamide gel electrophoresis under reducing or non-reducing conditions indicated that p45K has intramolecular but not intermolecular disulphide bonds, suggesting that interactions between p45K monomers may be due to some other type of association, such as hydrophobic or charge interactions. Velocity sedimentation of infected cell extracts revealed that native p45K also behaves as an oligomeric protein. Such results were confirmed using p45K partially purified by DEAE-cellulose chromatography. The evidence obtained indicated that all p45K present in the virion is in the oligomeric form, not associated by disulphide bonding, and that most native p45K present in the infected cells is also in the oligomeric form, probably as a consequence of early protein-protein interaction in rotavirus morphogenesis.