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Lytic Inactivation of Human Immunodeficiency Virus by Dual Engagement of gp120 and gp41 Domains in the Virus Env Protein Trimer.

ABSTRACT: We recently reported the discovery of a recombinant chimera, denoted DAVEI (dual-acting virucidal entry inhibitor), which is able to selectively cause specific and potent lytic inactivation of both pseudotyped and fully infectious human immunodeficiency virus (HIV-1) virions. The chimera is composed of the lectin cyanovirin-N (CVN) fused to the 20-residue membrane-proximal external region (MPER) of HIV-1 gp41. Because the Env gp120-binding CVN domain on its own is not lytic, we sought here to determine how the MPER(DAVEI) domain is able to endow the chimera with virolytic activity. We used a protein engineering strategy to identify molecular determinants of MPER(DAVEI) that are important for function. Recombinant mutagenesis and truncation demonstrated that the MPER(DAVEI) domain could be significantly minimized without loss of function. The dependence of lysis on specific MPER sequences of DAVEI, determination of minimal linker length, and competition by a simplified MPER surrogate peptide suggested that the MPER domain of DAVEI interacts with the Env spike trimer, likely with the gp41 region. This conclusion was further supported by observations from binding of the biotinylated MPER surrogate peptide to Env protein expressed on cells, monoclonal antibody competition, a direct binding enzyme-linked immunosorbent assay on viruses with varying numbers of trimeric spikes on their surfaces, and comparison of maximal interdomain spacing in DAVEI to that in high-resolution structures of Env. The finding that MPER(DAVEI) in CVN-MPER linker sequences can be minimized without loss of virolytic function provides an improved experimental path for constructing size-minimized DAVEI chimeras and molecular tools for determining how simultaneous engagement of gp120 and gp41 by these chimeras can disrupt the metastable virus Env spike.

SUBMITTER: Parajuli B 

PROVIDER: S-EPMC5444381 | BioStudies | 2016-01-01

REPOSITORIES: biostudies

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