Structral Study Of The Membrane-proximal Ectodomain Region Of HIV-1 Envelope Protein Gp41

Human immunodeficiency virus type 1 (HIV-1) is a T-cell lymphotropic virus, belonging to one of the retrovirus. The viruses spread wildly all over the world and threaten human life and health. HIV-1 mainly consists of three parts:envelope protein (Env), capsid and virus core. Env plays a crucial role in infection. It is initially produced as a precursor glycosylated gp160, which is processed by a host protease into two subunits:surface subunit gp120 and membrane-anchored subunit gp41. These two proteins are associated noncovalently and oligomerize as trimers on the surface of the virus. Only six broadly neutralizing monoclonal antibodies (bNtMAbs) have been isolated from human patient-derived material at present. Three of them recognize epitopes at gp120, while others,2F5,4E10 and Z13el, recognize conserved linear epitopes in the membrane proximal ectodomain region (MPER) of gp41.In this thesis, we studied the structure and location of gp41-MPER peptide and its L669A and L669S mutants in micelles by circular dichroism (CD) and nuclear magnetic resonance (NMR) spectral methods. At acidic pH values, the wild peptide (WT) in DPC micelles forms the L-shape structure with two discrete helical segments separated by a central hinge. The epitope of bNtMAbs 2F5 folds as an a-helix, while only half of the residues of epitope 4E10 folds as an a-helix and other residues of the epitope are unfolded. At a pH close to physiological environment (pH6.5). the content of a-helix decreases and both epitopes 2F5 and 4E10 include part of random coil other than a-helix. The mutation from Leu669 to Ala or Ser has little effect on the structure of the peptide in DPC micelles at acidic pH. These two mutant peptides also adopt predominantly an a-helical structure at this condition. When the pH value was increased to 6.5, the structure of the mutant peptide L669S is still similar to that of the wild peptide, but the a-helix content of the mutant peptide L669A increases evidently. At these two different pH values, the structures of the two epitopes are not altered by the mutations. The paramagnetic probe experiments at acidic pH reveal that WT and L669S insert the interior of DPC micelles, while L669A locates at the headgroup region of the micelles. At pH6.5, the insertion of the peptides WT and L669S in the micelles is less deeply than at acidic pH. Whereas the N-terminal part residues of WT are exposed to water and the C-terminal part residues are embedded in the headgroup region of DPC micelles, the entire residues of the peptide L669S locates at the headgroup region of the micelles. The increase in pH value induces little change in the location of the N-terminal part of the peptide L669A in micelles, but allows the C-terminal part more deeply inserted in the hydrophobic core of the micelles. This indicates that both the structure and the position of the MPER peptide in DPC micelles are affected by the mutation of L669A, but not by the mutation of L669S.The findings in this work are significant for understanding the structures of gp41 epitops and uncovering the mechanism of the virus function, and may provide useful information for the development of HIV-1 vaccines.