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HIV-1 envelope protein gp41: an NMR study of dodecyl phosphocholine embedded gp41 reveals a dynamic prefusion intermediate conformation.

ABSTRACT: Human immunodeficiency viral (HIV-1) fusion is mediated by the viral envelope gp120/gp41 complex (ENVelope glycoprotein). After gp120 shedding, gp41 is exposed and elicits membrane fusion via a cascade of conformational changes. In contrast to prefusion and postfusion conformation, little is known about any intermediate conformation. We report on a solution NMR investigation of homotrimeric HIV-1 gp41(27-194), comprising the transmembrane region and reconstituted in dodecyl phosphocholine (DPC) micelles. The protein is mainly ?-helical, but experiences internal dynamics on the nanosecond and micro to millisecond time scale and transient ?-helical behavior for certain residues in the N-terminal heptad repeat (NHR). Strong lipid interactions are observed, in particular for C-terminal residues of the NHR and imunodominant loop region connecting NHR and C-terminal heptad repeat (CHR). Our data indicate an extended conformation with features anticipated for a prefusion intermediate, presumably in exchange with a lowly populated postfusion six-helical bundle conformation.

SUBMITTER: Lakomek NA

PROVIDER: S-EPMC4156547 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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HIV-1 envelope protein gp41: an NMR study of dodecyl phosphocholine embedded gp41 reveals a dynamic prefusion intermediate conformation.

Lakomek Nils-Alexander NA Kaufman Joshua D JD Stahl Stephen J SJ Wingfield Paul T PT

Structure (London, England : 1993) 20140814 9

Human immunodeficiency viral (HIV-1) fusion is mediated by the viral envelope gp120/gp41 complex (ENVelope glycoprotein). After gp120 shedding, gp41 is exposed and elicits membrane fusion via a cascade of conformational changes. In contrast to prefusion and postfusion conformation, little is known about any intermediate conformation. We report on a solution NMR investigation of homotrimeric HIV-1 gp41(27-194), comprising the transmembrane region and reconstituted in dodecyl phosphocholine (DPC) ...[more]

PMID: 25132083

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Project description:Entry of human immunodeficiency virus type 1 (HIV-1) into host cells is mediated by conformational changes in the envelope glycoprotein (Env) that are triggered by Env binding to cellular CD4 and chemokine receptors. These conformational changes involve the opening of the gp120 surface subunit, exposure of the fusion peptide in the gp41 transmembrane subunit, and refolding of the gp41 N- and C-terminal heptad repeat regions (HR1 and HR2) first into an extended prehairpin intermediate and then into a compact 6-helix bundle (6HB) that facilitates fusion between viral and host cell membranes. Previously, we reported that Envs resistant to HR1 peptide fusion inhibitors acquired key resistance mutations in either HR1 or HR2 that increased 6HB stability. Here, we identify residues in HR1 that contribute not only to fusion inhibitor resistance and 6HB stability but also to reduced reactivity to CD4-induced conformational changes that lead to 6HB formation. While all Envs show increased neutralization sensitivity to mimetic CD4 (mCD4), Envs with either the E560K or Q577R HR1 mutation reduced conformational reactivity to CD4 that resisted viral inactivation and triggering to the 6HB. Using a panel of monoclonal antibodies (mAbs), we further determined that Envs from both HR1 and HR2 resistance pathways exhibit a relaxed trimer conformation due to gp120 adaptive mutations in different regions of Env that segregate by resistance pathway. These findings highlight regions of cross talk between gp120 and gp41 and identify HR1 residues that play important roles in regulating CD4-induced conformational changes in Env.IMPORTANCE Binding of the HIV envelope glycoprotein (Env) to cellular CD4 and chemokine receptors triggers conformational changes in Env that mediate virus entry, but premature triggering of Env conformational changes leads to virus inactivation. Currently, we have a limited understanding of the network of residues that regulate Env conformational changes. Here, we identify residues in HR1 of gp41 that modulate conformational changes in response to gp120 binding to CD4 and show that the mutations in HR1 and HR2 that confer resistance to fusion inhibitors are associated with gp120 mutations in different regions of Env that confer a more open conformation. These findings contribute to our understanding of the regulation of Env conformational changes and efforts to design new entry inhibitors and stable Env vaccine immunogens.

Project description:The mature human immunodeficiency virus (HIV-1) envelope glycoprotein (Env) trimer is produced by proteolytic cleavage of a precursor and consists of three gp120 exterior and three gp41 transmembrane subunits. The metastable Env complex is induced to undergo conformational changes required for virus entry by the binding of gp120 to the receptors, CD4 and CCR5/CXCR4. An isoleucine-to-proline change (I559P) in the gp41 ectodomain has been used to stabilize soluble forms of HIV-1 Env trimers for structural characterization and for use as immunogens. In the native membrane-anchored HIV-1BG505 Env, the I559P change modestly decreased proteolytic maturation, increased the non-covalent association of gp120 with the Env trimer, and resulted in an Env conformation distinctly different from that of the wild-type HIV-1BG505 Env. Compared with the wild-type Env, the I559P Env was recognized inefficiently by polyclonal sera from HIV-1-infected individuals, by several gp41-directed antibodies, by some antibodies against the CD4-binding site of gp120, and by antibodies that preferentially recognize the CD4-bound Env. Some of the gp120-associated antigenic differences between the wild-type HIV-1BG505 Env and the I559P mutant were compensated by the SOS disulfide bond between gp120 and gp41, which has been used to stabilize cleaved soluble Env trimers. Nonetheless, regardless of the presence of the SOS changes, Envs with proline 559 were recognized less efficiently than Envs with isoleucine 559 by the VRC01 neutralizing antibody, which binds the CD4-binding site of gp120, and the PGT151 neutralizing antibody, which binds a hybrid gp120-gp41 epitope. The I559P change completely eliminated the ability of the HIV-1BG505 Env to mediate cell-cell fusion and virus entry, and abolished the capacity of the SOS Env to support virus infection in the presence of a reducing agent. These results suggest that differences exist between the quaternary structures of functional Env spikes and I559P Envs.

Dataset Information

HIV-1 envelope protein gp41: an NMR study of dodecyl phosphocholine embedded gp41 reveals a dynamic prefusion intermediate conformation.

Publications

HIV-1 envelope protein gp41: an NMR study of dodecyl phosphocholine embedded gp41 reveals a dynamic prefusion intermediate conformation.

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