Human immunodeficiency virus type 1 capsid formation in reticulocyte lysates

Paul Spearman, Lee Ratner

Research output: Contribution to journalArticlepeer-review

40 Scopus citations

Abstract

The Gag polyprotein of human immunodeficiency virus (HIV) (Pr55(Gag)) contains sufficient information to direct particle assembly events when expressed within tissue culture cells. HIV Gag proteins normally form particles t a plasma membrane assembly site, in a manner analogous to that of the type C avian and mammalian leukemia/sarcoma viruses. It has not previously been demonstrated that immature HIV capsid can form without budding through an intact cellular membrane. In this study, a rabbit reticulocyte lysate translation reaction was used to recreate HIV capsid formation in vitro. Production of HIV-1 Pr55(Gag) and of a matrix-deleted Gag construct resulted in the formation of a subset of Gag protein structures with an equilibrium density of 1.15 g/ml. Gel filtration chromatography revealed these Gag protein structures to be larger than 2 x 106 Da, consistent with the formation of large multimers or capsid. These Gag protein structures were protease sensitive in the absence of detergent, indicating that did not a complete lipid envelope. Spherical structures were detected by electron microscopy within the reticulocyte lysate reaction mixtures and appeared and appeared essentially identical to immature HIV capsid or retrovirus-like particles. These results demonstrate that the HIV Gag protein is capable of producing immature capsid in a cell-free reaction and that such capsid lack as complete lipid envelope.

Original languageEnglish
Pages (from-to)8187-8194
Number of pages8
JournalJournal of virology
Volume70
Issue number11
DOIs
StatePublished - Nov 1996

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