Alanine Scanning Mutants of the HIV gp41 Loop

Lijun Rong,Amy Jacobs,Michael Caffrey,Jayita Sen

doi:10.1074/jbc.m414411200

Lijun Rong, Amy Jacobs + Show 2 more

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https://doi.org/10.1074/jbc.m414411200

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Abstract

Based on mutagenesis and structural studies of human immunodeficiency virus (HIV) envelope proteins, the loop region of gp41 is thought to directly interact with gp120. The importance of the HIV gp41 loop region to envelope function has been systematically examined by alanine scanning of all gp41 loop residues and the subsequent characterization of the mutagenic effects on viral entry, envelope expression, envelope processing, and gp120 association with gp41. With respect to the wild-type gp41, mutational effects on viral entry fall into four classes as follows: 1) little or no effect (G594A, S599A, G600A, K601A, N611A, S615A, N616A, and L619A); 2) significantly reduced entry (I595A, L602A, I603A, V608A, and K617A); 3) abolished entry (L593A, W596A, G597A, T606A, W610A, W614A, S618A, and I622A); and 4) enhanced entry (T605A, P609A, S613A, E620A, and Q621A). The reduced functionality of many mutants was apparently due to either disruption of envelope processing (L593A and T606A), viral incorporation of the envelope (W610A, W614A, and I662A), or increased dissociation of gp120 (W596A, G597A, and S618A). The extreme sensitivity of the gp120-gp41 interaction to alanine substitutions (e.g. the G597A and S618A mutants are relatively conservative substitutions) suggests that this association is an attractive and novel target for future drug discovery efforts.

Highlights

Infection with the human immunodeficiency virus (HIV)1 begins when the virus particle attaches to the cell surface and the viral membrane fuses with the target cell membrane, thereby allowing entry of the viral genetic material
Many of the alanine substitutions are relatively conservative in nature (e.g. G597A, T606A, and S618A)
Trp614 occurs within the four-residue insertion of HIV-1 gp41 with respect to HIV-2 and simian immunodeficiency virus (SIV) (Fig. 1), which suggests that this region plays a critical role in HIV-1 gp41 function

Summary

Introduction

Infection with the human immunodeficiency virus (HIV) begins when the virus particle attaches to the cell surface and the viral membrane fuses with the target cell membrane, thereby allowing entry of the viral genetic material (reviewed in Ref. 1). The viral envelope proteins gp120 and gp, which form a non-covalent complex on the viral surface, play critical roles in viral attachment and membrane fusion [2]. There is extensive information on the extracellular domains, or ectodomains, of HIV and SIV gp from x-ray, NMR, and molecular modeling studies [5,6,7,8,9,10,11,12]. There is no structural information available for the gp120-gp complex; based on previous mutagenesis studies, the central loop of gp is one site of non-covalent interactions with gp120 that are critical to HIV entry [13,14,15,16,17]. We present the first systematic mutation study of the importance of each gp loop residue to gp function with the long-term goal of identifying gp regions that are attractive sites for drug intervention

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