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Lytic Inactivation of HIV-1 by Dual Engagement of gp120 and gp41 Domains in the Virus Env Protein Trimer.

Lytic Inactivation of HIV-1 by Dual Engagement of gp120 and gp41 Domains in the Virus Env Protein Trimer.
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Parajuli B, Acharya K, Yu R, Ngo B, Rashad AA, Abrams CF, Chaiken IM,


Parajuli B, Acharya K, Yu R, Ngo B, Rashad AA, Abrams CF, Chaiken IM, (click to view)

Parajuli B, Acharya K, Yu R, Ngo B, Rashad AA, Abrams CF, Chaiken IM,

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Biochemistry 2016 Oct 12()

Abstract

We recently reported 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 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. Since 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) important for function. Recombinant mutagenesis and truncation demonstrated that the MPER(DAVEI) domain could be significantly minimized without loss of function. 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 biotinylated MPER surrogate peptide to Env protein expressed on cells, monoclonal antibody competition, direct binding ELISA on viruses with varying number of trimeric spikes on their surfaces and comparison of maximum inter-domain spacing in DAVEI to 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 to construct size-minimized DAVEI chimeras and molecular tools to determine how simultaneous engagement of gp120 and gp41 by these chimeras can disrupt the metastable virus Env spike.

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