Efficient production of HIV-1 virus-like particles from a mammalian expression vector requires the N-terminal capsid domain
It is now well accepted that the structural protein Pr55(Gag) is sufficient by itself to produce HIV-1 virus-like particles (VLPs). This polyprotein precursor contains different domains including matrix, capsid, SP1, nucleocapsid, SP2 and p6. In the present study, we wanted to determine by mutagenes...
Saved in:
Published in | PloS one Vol. 6; no. 11; p. e28314 |
---|---|
Main Authors | , , , , |
Format | Journal Article |
Language | English |
Published |
United States
Public Library of Science
30.11.2011
Public Library of Science (PLoS) |
Subjects | |
Online Access | Get full text |
Cover
Loading…
Summary: | It is now well accepted that the structural protein Pr55(Gag) is sufficient by itself to produce HIV-1 virus-like particles (VLPs). This polyprotein precursor contains different domains including matrix, capsid, SP1, nucleocapsid, SP2 and p6. In the present study, we wanted to determine by mutagenesis which region(s) is essential to the production of VLPs when Pr55(Gag) is inserted in a mammalian expression vector, which allows studying the protein of interest in the absence of other viral proteins. To do so, we first studied a minimal Pr55(Gag) sequence called Gag min that was used previously. We found that Gag min fails to produce VLPs when expressed in an expression vector instead of within a molecular clone. This failure occurs early in the cell at the assembly of viral proteins. We then generated a series of deletion and substitution mutants, and examined their ability to produce VLPs by combining biochemical and microscopic approaches. We demonstrate that the matrix region is not necessary, but that the efficiency of VLP production depends strongly on the presence of its basic region. Moreover, the presence of the N-terminal domain of capsid is required for VLP production when Gag is expressed alone. These findings, combined with previous observations indicating that HIV-1 Pr55(Gag)-derived VLPs act as potent stimulators of innate and acquired immunity, make the use of this strategy worth considering for vaccine development. |
---|---|
Bibliography: | Conceived and designed the experiments: PJ KT AD RC MJT. Performed the experiments: PJ KT AD. Analyzed the data: PJ KT AD RC MJT. Contributed reagents/materials/analysis tools: PJ KT. Wrote the paper: PJ RC MJT. |
ISSN: | 1932-6203 1932-6203 |
DOI: | 10.1371/journal.pone.0028314 |