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Principal Investigator: Adriaan Bax
Organization: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
Fiscal Year: 2021
Award: $658,712
Funding agency: National Institute of Diabetes and Digestive and Kidney Diseases
The envelope glycoprotein gp41 mediates the process of membrane fusion that enables entry of the HIV-1 virus into the host cell. On the surface of the virus, gp41 is part of the large homotrimeric glycoprotein gp160, and previous work by us has shown that, in contrast with literature reports, the transmembrane helical anchor of gp41, which spans the viral membrane, does not significantly contribute to its trimerization propensity. We expanded our structural study to the membrane-proximal external region (MPER) of HIV-1 gp41, which contains epitopes for at least four broadly neutralizing antibodies. In prior studies, depending on solution conditions and construct design, different structures have been reported for this segment. Our new results found that in aqueous solution the MPER fragment (gp160 residues 660-674) exists in a monomer-trimer equilibrium with an association constant in the micro-molar range. Thermodynamic analysis revealed that the association is exothermic, more favorable in D2O than H2O, and increased with ionic strength, indicating hydrophobically driven intermolecular interactions. Circular dichroism, 13C chemical shifts, NOE, and hydrogen exchange rates revealed that MPER undergoes a structural transition from predominately unfolded monomer at low concentrations to an alpha-helical trimer at high concentrations. This result has implications for antibody recognition of MPER prior to and during the process where gp41 switches from a pre-hairpin intermediate to its post-fusion 6-helical bundle state.
Preliminary experiments applied to the ecto-domain of the Spike protein of SARS-CoV-2 indicate this protein contains a three-helical N-terminal heptad repeat (NHR) that is considerably more stable than the corresponding region of gp41, and the propensity for the C-terminal heptad (CHR) repeat albeit lower than that of NHR, is also substantially higher than for the CHR of gp41, whereas the propensity to trimerize for its membrane proximal region is substantially lower.
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