High efficient production of Pr55(gag) virus-like particles expressing multiple HIV-1 epitopes, including a gp120 protein derived from an Ugandan HIV-1 isolate of subtype A

The main goal of this study was to investigate a novel approach for an effi cient and reproducible production of Virus-Like Particles (VLPs) expressing multiple HIV-1 epitopes. The HIV-1 Pr55(gag)-based VLPs have been produced in a Baculovirus expression system, using a transfer vector able to suppor t the independent expression of different open reading frames (ORFs). In th is regard, the gp120 derived from 94UG018 HIV-1(A) isolate, previously stud ied in our laboratory, has been packaged into the VLPs together with nef an d pol ORFs. In particular. the gp120(UG) sequence shows a 90% homology in t he V3 region competed to African HIV-1 strains of the A-clade. This novel a pproach is extremely effective for the production of VLPs expressing all th e epitopes, as confirmed by Western Blot characterization. Furthermore, the resulting HIV-VLP(A)s show the expected density(1.14-1.18 g/ml on a 10-60% sucrose gradient and the morphology of an immature virion at standard tran smission electron microscopy. Our results demonstrate that this strategy is highly efficient for expressing a balanced amount of multiple epitopes and their packaging in VLP structures, without affecting the Pr55(gag) autoass embling capacities. Furthermore, the genetic transposition performed in a m odified E. coli represents a methodological improvement. allowing a faster and more reproducible identification of recombinant Baculovirus DNA molecul es. (C) 2001 Elsevier Science B.V. All rights reserved.