STABILITY AND PROTEOLYTIC DOMAINS OF NEF PROTEIN FROM HUMAN-IMMUNODEFICIENCY-VIRUS (HIV) TYPE-1

Proteolytic experiments in conjunction with H-1-NMR spectroscopy show that the Nef (negative factor) protein from human immunodeficiency vir us type 1 probably consists of two main domains, the N-terminal anchor domain at amino acid positions 2-65 and the C-terminal core domain at positions 66-206. The N-terminal domain is likely to be located at th e surface of the protein, while the C-terminal domain has a compactly folded core and is stable in the absence of the anchor domain. It is c onceivable that the core domain represents a functional domain of the Nef protein, activated after the removal of the membrane anchor by the human-immunodeficiency-virus protease or cellular proteases. Nef is s table at pH 5-12 and denatures at 317-322 K. The Nef protein remains i n its native conformation in dimethyl-sulfoxide/water mixtures up to 3 5% (by vol.), and in acetonitrile/water up to 14% (by vol.). Nef refol ds spontaneously after denaturation with urea or guanidinium hydrochlo ride. The H-1-NMR parameters and pK(a) values of five of the nine hist idine residues and one of the seven tyrosine residues were determined and were found in four cases to be typical for residues which are not located in the interior of the protein.