CHARACTERIZATION OF P91, A PROTEIN ASSOCIATED WITH VIRIONS OF AN ORGYIA-PSEUDOTSUGATA BACULOVIRUS

Polyclonal antiserum produced against preoccluded virions from the Org yia pseudotsugata multinucleocapsid nuclear polyhedrosis virus (OpMNPV ) was used to screen an OpMNPV lambda gt11 expression library. The ins ert from one of the immunoreactive phage isolates hybridized to OpMNPV orf86 (p91), a 2460-bp (819 amino acids) open reading frame that enco des a predicted protein of 91 kDa. Antibodies generated against a malt ose binding protein-P91 fusion detected a band of approximately 91 kDa on Western blots of extracts of OpMNPV-infected Lymantria dispar cell s. This band was first observed al 18 hr p.i, and was present at all l ater time points. Similar results using this antiserum were seen with a time course of Autographa californica-infected Spodoptera frugiperda cells. Localization of P91 by confocal immunofluorescence microscopy showed that the protein was concentrated near the nuclear membrane and at late times p.i. was most concentrated near polyhedra. Immunoelectr on microscopy indicated that P91 was present in both the capsid and en velope surrounding the capsid of occlusion-derived virions. Fractionat ion studies employing NP-40 and Western blot analysis indicated that P 91 was associated with the capsid structure. (C) 1997 Academic Press.