Characterisation of the redox changes in the HIV envelope glycoprotein during cell entry

A functional disulfide bond in both the HIV envelope glycoprotein, gp120, and its immune cell receptor, CD4, are involved in viral entry and compounds that block cleavage of the disulfide bond in these proteins inhibit HIV entry and infection. The disulfide bonds in both proteins are cleaved at the cell surface by the small redox protein, thioredoxin. A thioredoxin kinetic trapping mutant cleaved a single disulfide bond in isolated and cell surface gp120, but not the gp160 precursor, and the extent of the reaction was enhanced when gp120 was bound to CD4. The target gp120 disulfide and its mechanism of cleavage were determined by mass spectrometry analysis of the gp120-thioredoxin complex. The Cys32 sulfur ion of thioredoxin attacks the Cys296 sulfur ion of the gp120 V3 domain Cys296-Cys331 disulfide bond, cleaving the bond. There are 20 possible disulfide bond configurations and, notably, the V3 domain disulfide has the same unusual -RHStaple configuration as the Cys130-Cys159 disulfide bond cleaved in CD4. Considering that V3 sequences largely determine the chemokine receptor preference of HIV we propose that cleavage of the V3 domain disulfide, which is facilitated by CD4 binding, regulates chemokine receptor binding. A conformational change in the V3 domain as a result of cleavage of the Cys296-Cys331 bond may result in dissociation of chemokine receptor from gp120, which allows refolding of gp41 and viralmembrane fusion. Strategies to prevent cleavage of the gp120 V3 disulfide bond or the conformational change in the domain following cleavage will inhibit HIV infection.