title: A Highly Conserved gp120 Inner Domain Residue Modulates Env Conformation and Trimer Stability.
abstract: Previous studies have shown that highly conserved residues in the inner domain of gp120 are required for HIV-1 envelope glycoprotein (Env) transitions to the CD4-bound conformation (A. Finzi, S. H. Xiang, B. Pacheco, L. Wang, J. Haight, et al., Mol Cell 37:656-667, 2010, http://dx.doi.org/10.1016/j.molcel.2010.02.012; A. Desormeaux, M. Coutu, H. Medjahed, B. Pacheco, A. Herschhorn, et al., J Virol 87:2549-2562, 2013, http://dx.doi.org/10.1128/JVI.03104-12). Moreover, W69, a highly conserved residue located at the interface between layer 1 and layer 2 of the inner domain, was recently shown to be important for efficient Env recognition by CD4-induced (CD4i) antibodies capable of potent antibody-dependent cellular cytotoxicity (W. D. Tolbert, N. Gohain, M. Veillette, J. P. Chapleau, C. Orlandi, et al., 2016, Structure 24:697-709, http://dx.doi.org/10.1016/j.str.2016.03.005; S. Ding, M. Veillette, M. Coutu, J. Prevost, L. Scharf, et al., 2016, J Virol 90:2127-2134, http://dx.doi.org/10.1128/JVI.02779-15). We evaluated the contribution of the hydrophobicity of W69 to conformational changes of Env by replacing it with a series of residues with aliphatic or aromatic side chains of decreasing chain length. We have found that the hydrophobicity of residue 69 is important for Env processing, CD4 binding, and its transition to the CD4-bound conformation. The most deleterious effect was observed when W69 was replaced with alanine or glycine residues. However, the functions lost due to W69 mutations could be progressively restored with amino acids of increasing aliphatic chain length and fully recovered with residues bearing an aromatic ring. Interestingly, poor CD4 binding of W69A could be fully restored by introducing a compensatory mutation within layer 2 (S115W). Structural studies of HIV-1 gp120 coree W69A/S115W mutant bound to the CD4 peptide mimetic M48U1 and Fab of anti-cluster A antibody N60-i3 revealed no perturbations to the overall structure of the double mutant compared to the wild-type protein but identified higher mobility within the interface between layer 1 and layer 2, the bridging sheet region, and the CD4 binding site.IMPORTANCE HIV-1 Env transitions to the CD4-bound conformation are required for viral entry. Previous studies identified a highly conserved residue of the inner domain, W69, as being involved in these conformational transitions (A. Finzi, S. H. Xiang, B. Pacheco, L. Wang, J. Haight, et al., Mol Cell 37:656-667, 2010, http://dx.doi.org/10.1016/j.molcel.2010.02.012). Here, we show that W69, located at the interface between gp120 and gp41 in the PGT151-bound trimer, plays a critical role in the interprotomer signaling induced by CD4 binding. This new information might be useful in immunogen design.
authors.affiliation: Centre de Recherche du CHUM, Université de Montréal, Montreal, Quebec, Canada Department of Microbiology, Infectiology and Immunology, Université de Montréal, Montreal, Quebec, Canada.
authors.forename: Shilei
authors.initials: S
authors.lastname: Ding
authors.affiliation: Institute of Human Virology and Department of Biochemistry and Molecular Biology, University of Maryland, Baltimore, Maryland, USA.
authors.forename: William D
authors.initials: WD
authors.lastname: Tolbert
authors.affiliation: Centre de Recherche du CHUM, Université de Montréal, Montreal, Quebec, Canada Department of Microbiology, Infectiology and Immunology, Université de Montréal, Montreal, Quebec, Canada.
authors.forename: Jérémie
authors.initials: J
authors.lastname: Prévost
authors.affiliation: Centre de Recherche du CHUM, Université de Montréal, Montreal, Quebec, Canada.
authors.forename: Beatriz
authors.initials: B
authors.lastname: Pacheco
authors.affiliation: Centre de Recherche du CHUM, Université de Montréal, Montreal, Quebec, Canada Department of Microbiology, Infectiology and Immunology, Université de Montréal, Montreal, Quebec, Canada.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Coutu
authors.affiliation: Centre de Recherche du CHUM, Université de Montréal, Montreal, Quebec, Canada.
authors.forename: Olfa
authors.initials: O
authors.lastname: Debbeche
authors.affiliation: University of Nebraska, Lincoln, Nebraska, USA.
authors.forename: Shi-Hua
authors.initials: SH
authors.lastname: Xiang
authors.affiliation: Institute of Human Virology and Department of Biochemistry and Molecular Biology, University of Maryland, Baltimore, Maryland, USA andres.finzi@umontreal.ca mpazgier@ihv.umaryland.edu.
authors.forename: Marzena
authors.initials: M
authors.lastname: Pazgier
authors.affiliation: Centre de Recherche du CHUM, Université de Montréal, Montreal, Quebec, Canada Department of Microbiology, Infectiology and Immunology, Université de Montréal, Montreal, Quebec, Canada Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada andres.finzi@umontreal.ca mpazgier@ihv.umaryland.edu.
authors.forename: Andrés
authors.initials: A
authors.lastname: Finzi
mesh_terms: D019943:Amino Acid Substitution; D017124:Conserved Sequence; D004252:DNA Mutational Analysis; D015699:HIV Envelope Protein gp120 / Q000737:chemistry* / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D055550:Protein Stability
chemical_list: D015699:HIV Envelope Protein gp120; C514793:gp120 protein, Human immunodeficiency virus 1