Cross-functional analysis of the Microviridae internal scaffolding protein

The assembly of the viral structural proteins into infectious virions is of ten mediated by scaffolding proteins. These proteins are transiently associ ated with morphogenetic intermediates but not found in the mature particle. The genes encoding three Microviridae (empty setX174, G4 and alpha 3) inte rnal scaffolding proteins (B proteins) have been cloned, expressed in vivo and assayed for the ability to complement null mutations of different Micro viridae species. Despite divergence as great as 70% in amino acid sequence over the aligned length, cross-complementation was observed, indicating tha t these proteins are capable of directing the assembly of foreign structura l proteins into infectious particles. These results suggest that the Microv iridae internal scaffolding proteins may be inherently flexible. There was one condition in which a B protein could not cross-function. The empty setX 174 B protein cannot productively direct the assembly of the G4 capsid at t emperatures above 21 degrees C. Under these conditions, assembly is arreste d early in the morphogenetic pathway, before the first B protein mediated r eaction. Two G4 mutants, which can productively utilize the empty setX174 B protein at elevated temperatures, were isolated. Both mutations confer ami no acid substitutions in the viral coat protein but differ in their relativ e abilities to utilize the foreign scaffolding protein. The more efficient substitution is located in a region where coat-scaffolding interactions hav e been observed in the atomic structure and may emphasize the importance of interactions in this region. (C) 1999 Academic Press.