Functional links between the fusion peptide-proximal polar segment and membrane-proximal region of human immunodeficiency virus gp41 in distinct phases of membrane fusion

Anna Kathryn Bellamy-McIntyre, Chan-Sien Lay, Severine Bar, Anne L Maerz, Gert Hoy Talbo, Heidi Drummer, Pantelis Poumbourios

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60 Citations (Scopus)

Abstract

The binding of CD4 and chemokine receptors to the gp120 attachment glycoprotein of human immunodeficiency virus triggers refolding of the associated gp41 fusion glycoprotein into a trimer of hairpins with a 6-helix bundle (6HB) core. These events lead to membrane fusion and viral entry. Here, we examined the functions of the fusion peptide-proximal polar segment and membrane-proximal Trp-rich region (MPR), which are exterior to the 6HB. Alanine substitution of Trp(666), Trp(672), Phe(673), and Ile(675) in the MPR reduced entry by up to 120-fold without affecting gp120-gp41 association or cell-cell fusion. The L537A polar segment mutation led to the loss of gp120 from the gp120-gp41 complex, reduced entry by approximately 10-fold, but did not affect cell-cell fusion. Simultaneous Ala substitution of Leu(537) with Trp(666), Trp(672), Phe(673), or Ile(675) abolished entry with 50-80 reductions in cell-cell fusion. gp120-gp41 complexes of fusion-defective double mutants were resistant to soluble CD4-induced shedding of gp120, suggesting that their ability to undergo receptor-induced conformational changes was compromised. Consistent with this idea, a representative mutation, L537A/W666A, led to an approximately 80 reduction in lipophilic fluorescent dye transfer between gp120-gp41-expressing cells and receptor-expressing targets, indicating a block prior to the lipid-mixing phase. The L537A/W666A double mutation increased the chymotrypsin sensitivity of the polar segment in a trimer of hairpins model, comprising the 6HB core, the polar segment, and MPR linked N-terminally to maltose-binding protein. The data indicate that the polar segment and MPR of gp41 act synergistically in forming a fusion-competent gp120-gp41 complex and in stabilizing the membrane-interactive end of the trimer of hairpins.
Original languageEnglish
Pages (from-to)23104 - 23116
Number of pages13
JournalThe Journal of Biological Chemistry
Volume282
Issue number32
Publication statusPublished - 2007

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