The methods developed in the course of our earlier work with pyruvate dehydrogenase has served as a platform for studies on HIV and SIV envelope glycoproteins and have resulted in a number of new structures. These studies, which produced the first structures of trimeric Env immunogens a couple of years ago, are now leading to new insights that could be invaluable for rational vaccine design. Two examples are listed below. In one instance, we reported cryo-electron microscopic studies of the interaction between the ectodomain of trimeric HIV-1 envelope glycoprotein (Env) and Z13e1, a broadly neutralizing antibody that targets the membrane proximal external region (MPER) of the gp41 subunit. We showed that Z13e1-bound Env displays an open quaternary conformation similar to the CD4-bound conformation. Our results support the idea that MPER directed antibodies, such as Z13e1, block viral entry by interacting with Env at a step after CD4 activation. In another example, we carried out cryo-electron microscopic analysis at sub-nanometer resolution of a cleaved, soluble version of trimeric Env to explore structural changes that take place with ligand activation. We showed that Env in the open, activated conformation has highly conserved elements of gp41 organized in a three-helix motif in the central portion of the envelope glycoprotein complex. The N-terminal gp41 helices in this novel, activated Env conformation are much less compactly packed than in the post-fusion, six-helix bundle state, suggesting a new structural template for designing immunogens that can elicit antibodies targeting HIV at a vulnerable, pre-entry stage.

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Harris, Audray K; Bartesaghi, Alberto; Milne, Jacqueline L S et al. (2013) HIV-1 envelope glycoprotein trimers display open quaternary conformation when bound to the gp41 membrane-proximal external-region-directed broadly neutralizing antibody Z13e1. J Virol 87:7191-6
Lengyel, Jeffrey S; Stott, Katherine M; Wu, Xiongwu et al. (2008) Extended polypeptide linkers establish the spatial architecture of a pyruvate dehydrogenase multienzyme complex. Structure 16:93-103
Liu, Jun; Bartesaghi, Alberto; Borgnia, Mario J et al. (2008) Molecular architecture of native HIV-1 gp120 trimers. Nature 455:109-13
Lengyel, Jeffrey S; Milne, Jacqueline L S; Subramaniam, Sriram (2008) Electron tomography in nanoparticle imaging and analysis. Nanomedicine (Lond) 3:125-31
Nanavati, Dhaval; Gucek, Marjan; Milne, Jacqueline L S et al. (2008) Stoichiometry and absolute quantification of proteins with mass spectrometry using fluorescent and isotope-labeled concatenated peptide standards. Mol Cell Proteomics 7:442-7