[MoOrA1093] Creation of cleaved, stabilized HIV-1 envelope glycoprotein trimers

14th International AIDS Conference

Barcelona, Spain — July 7-12, 2002

[TITLE:] Creation of cleaved, stabilized HIV-1 envelope glycoprotein trimers

[AUTHOR(S):] J.P. Moore, R.W. Sanders, M. Vesanen, S. Beddows¹, M. Lu², N. Schuelke, J. Gardner, P.J. Maddon, W.C. Olson³

Int Conf AIDS. 2002 Jul 7-12;14:Abstract No. MoOrA1093

[ABSTRACT:] The native HIV-1 envelope complex is a trimeric structure containing 3 gp120 and 3 gp41 subunits, all associated via non-covalent interactions. This complex is the target for neutralizing antibodies, and may be an effective immunogen for inducing these antibodies. Unfortunately, the native complex is too unstable to be made in quantity as a recombinant protein. To address this problem, we have been seeking ways to stabilize the inter-subunit interactions within the complex. These proteins are expressed as soluble gp140s, because a stop-codon has been introduced immediately before the transmembrane domain of gp41. The association between the gp120 and gp41 subunits in the gp140 proteins can be stabilized by the introduction of appropriately positioned cysteine residues that create an intermolecular disulfide bond. These proteins, SOS gp140, are fully cleaved by furin proteases. However, the SOS gp140 complex is still unstable, and is mostly monomeric. Thus, SOS gp140 trimers cannot be purified. In contrast, uncleaved gp140 proteins form dimers, trimers and tetramers, implying that Env instability is caused by gp140 cleavage into gp120-gp41. We have now stabilized the SOS gp140 protein in its trimeric, pre-fusion form by introducing an amino acid substitution into gp41 that disfavors the transition to the 6-helix bundle, post-fusion form of the protein. This protein, SOSIP gp140, is stable, fully cleaved and trimeric. It can be converted to a gp140 monomer by mild heat (>50 C) or by SDS, but is stable to high salt, non-ionic detergent and freeze-thaw cycles. Stabilized, cleaved trimers can also be made by removing the V1V2 loop structure from SOS gp140. The antigenic and receptor-binding properties of purified SOSIP gp140 trimers are now being compared with those of delta-loop SOS gp140 trimers, and uncleaved gp140 dimers, trimers and tetramers. Investigations of the immunogenicity of SOSIP gp140 and delta-loop SOS gp140 trimers are planned for early 2002.

Presenting author: John Moore

1Weill Medical College of Cornell University, Dept. of Microbiology & Immunology, Weill Medical College of Cornell University, 1300 York Avenue, Box 62, New York, NY 10021, United States.

2Weill Medical College of Cornell University, Dept. of Biochemistry, New York, United States.

3Progenics Pharmaceuticals, Tarrytown, United States.

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