Functional analysis of the disulfide-bonded loop/chain reversal region of human immunodeficiency virus type 1 gp41 reveals a critical role in gp120-gp41 association

Human immunodeficiency virus type 1 (HIV-1) entry into cells is mediated by the surface-exposed envelope protein (SU) gp120, which binds to cellular C D4 and chemokine receptors, triggering the membrane fusion activity of the transmembrane (TM) protein gp41. The core of gp41 comprises an N-terminal t riple-stranded coiled coil and an antiparallel C-terminal helical segment w hich is packed against the exterior of the coiled coil and is thought to co rrespond to a fusion-activated conformation. The available gp41 crystal str uctures lack the conserved disulfide-bonded loop region which, in human T l ymphotropic virus type 1 (HTLV-1) and murine leukemia virus TM proteins, me diates a chain reversal, connecting the antiparallel N- and C-terminal regi ons. Mutations in the HTLV-1 TM protein gp21 disulfide-bonded loop/chain re versal region adversely affected fusion activity without abolishing SU-TM a ssociation (A. L. Maerz, R. J. Center, B. E. Kemp, B. Kobe, and P. Poumbour ios, J, Virol. 74:6614-6621, 2000). We now report that in contrast to our f indings,vith HTLV-1, conservative substitutions in the HIV-1 gp41 disulfide -bonded loop/chain reversal region abolished association with gp120. While the mutations affecting gp120-gp41 association also affected cell-cell fusi on activity, HIV-1 glycoprotein maturation appeared normal. The mutant glyc oproteins were processed, expressed at the cell surface, and efficiently im munoprecipitated by conformation-dependent monoclonal antibodies. The gp120 association site includes aromatic and hydrophobic residues on either side of the gp41 disulfide-bonded loop and a basic residue within the loop. The HIV-1 gp41 disulfide-bonded loop/chain reversal region is a critical gp120 contact site; therefore, it is also likely to play a central role in fusio n activation by linking CD4 plus chemokine receptor-induced conformational changes in gp120 to gp41 fusogenicity. These gp120 contact residues are pre sent in diverse primate lentiviruses, suggesting conservation of function.