Involvement of the C-Terminal Disulfide-Bonded Loop of Murine Leukemia Virus SU Protein in a Postbinding Step Critical for Viral Entry

Abstract

A role for the C-terminal domain (CTD) of murine leukemia virus (MuLV) Env protein in viral fusion was indicated by the potent inhibition of MuLV-induced fusion, but not receptor binding, by two rat monoclonal antibodies (MAbs) specific for epitopes in the CTD. Although these two MAbs, 35/56 and 83A25, have very different patterns of reactivity with viral isolates, determinants of both epitopes were mapped to the last C-terminal disulfide-bonded loop of SU (loop 10), and residues in this loop responsible for the different specificities of these MAbs were identified. Both MAbs reacted with a minor fraction of a truncated SU fragment terminating four residues after loop 10, indicating that while the deleted C-terminal residues were not part of these epitopes, they promoted their formation. Neither MAb recognized the loop 10 region expressed in isolated form, suggesting that these epitopes were not completely localized within loop 10 but required additional sequences located N terminal to the loop. Direct support for a role for loop 10 in fusion was provided by the demonstration that Env mutants containing an extra serine or threonine residue between the second and third positions of the loop were highly attenuated for infectivity and defective in fusion assays, despite wild-type levels of expression, processing, and receptor binding. Other mutations at positions 1 to 3 of loop 10 inhibited processing of the gPr80 precursor protein or led to increased shedding of SU, suggesting that loop 10 also affects Env folding and the stability of the interaction between SU and TM.