Design and properties of N-CCG-gp41, a chimeric gp41 molecule with nanomolar HIV fusion inhibitory activity

The design and characterization of a chimeric protein, termed N-CCG-gp41, d erived from the ectodomain of human immunodeficiency virus (HIV), type I gp 41 is described. N-CCG-gp41 features an exposed trimeric coiled-coil compri sing the N-terminal helices of the gp41 ectodomain. The trimeric coiled-coi l is stabilized both by fusion to a minimal thermostable ectodomain of gp41 and by engineered intersubunit disulfide bonds. N(CCG)gp41 is shown to inh ibit HIV envelope-mediated cell fusion at nanomolar concentrations with an IC., of 16.1 +/- 2.8 nM. It is proposed that N-CCG-gp41 targets the exposed C-terminal region of the gp41 ectodomain in its pre-hairpin intermediate s tate, thereby preventing the formation of the fusogenic form of the gp41 ec todomain, which comprises a highly stable trimer of hairpins arranged in a six-helix bundle. N-CCG-gp41 has potential as a therapeutic agent for the d irect inhibition of HIV cell entry, as an anti-HIV vaccine, and as a compon ent of a rapid throughput assay for screening for small molecule inhibitors of HIV envelope-mediated cell fusion. It is anticipated that antibodies ra ised against N-CCG-gp41 may target the trimeric coiled-coil of N-terminal h elices of the gp41 ectodomain that is exposed in the pre-hairpin intermedia te state in a manner analogous to peptides derived from the C-terminal heli x of gp41 that are currently in clinical trials.