Expression of HIV-1 gag and pol Gene Products using Recombinant Baculoviruses
Since its original description some years ago (Smith et al., 1983a, 1983b), recombinant baculoviruses have become an increasingly popular method for the expression of heterologous gene products. Genes of interest are introduced into the genome of the Autographica callfornica nuclear polyhedrosis virus (AcNPV) to give a helper independant recombinant virus that, upon infection of insect cells, produces the product encoded by the introduced gene in a regulated manner. Frequently, though not always, excellent yields of recombinant protein are produced (for a review see Luckow and Summers 1988a). Moreover, as the host expression background is a higher eucaryotic cell, many post-translational modificaitions such as myristylation, phosphorylation and glycosylation are efficiently carried out, leading to recombinant products that are often indistinguishable from their ‘natural’ counterparts. We have used this system to produce some ten different recombinant baculoviruses expressing various gene products encoded by both HIV-1 and HIV-2. Here we describe four of these recombinant viruses that produce products encoded by the gag and pol genes of HIV-1. We show that a feature of gag and pol translation in HIV-1 infected cells, the ribosomal frame shift, also occurs in insect cells leading to production of the HIV protease and specific cleavage of the gag precursor. The gag product, p55, and the protease can also be produced independently using recombinant baculoviruses and when co-infected into insect cells, cleavage of p55 in trans is observed. In the absence of cleavage by the protease, the p55 gag precusor molecule assembles into structures closely resembling the HIV-1 pre-core. When only p24 is expressed however, and despite very high expression levels, no such structures are observed suggesting that the primary amino acid sequences necessary for subunit interaction and assembly do not lie within the p24 coding region.
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- Summers, M.D. and Smith, G.E. ‘A Manual of Methods for Baculovirus vectors and Insect Cell Culture Proceedures’, Texas Agricultural Experimental Station Bulletin No. 1555 (1987).Google Scholar