Delta-peptide is the carboxy-terminal cleavage fragment of the nonstructural small glycoprotein sGP of Ebola virus

In the present study we have investigated processing and maturation of the nonstructural small glycoprotein (sGP) of Ebola virus. When sGP expressed f rom vaccinia virus vectors was analyzed by pulse-chase experiments using SD S-PAGE under reducing conditions, the mature form and two different precurs ors have been identified. First, the endoplasmic reticulum form sGP(er), fu ll-length sGP with oligomannosidic N-glycans, was detected, sGP(or) was the n replaced by the Golgi-specific precursor pre-sGP, full-length sGP contain ing complex N-glycans. This precursor was finally converted by proteolysis into mature sGP and a smaller cleavage fragment, Delta-peptide. Studies emp loying site-directed mutagenesis revealed that sGP was cleaved at a multiba sic amino acid motif at positions 321 to 324 of the open reading frame. Cle avage was blocked by RVKR-chloromethyl ketone. Uncleaved pre-sGP forms a di sulfide-linked homodimer and is secreted into the culture medium in the pre sence of the inhibitor as efficiently as proteolytically processed sGP. In vitro treatment of pre-sGP by purified recombinant furin resulted in effici ent cleavage, confirming the importance of this proprotein convertase for t he processing and maturation of sGP. Delta-peptide is also secreted into th e culture medium and therefore represents a novel nonstructural expression product of the GP gene of Ebola virus. Both cleavage fragments contain sial ic acid, but only Delta-peptide is highly O-glycosylated. (C) 1999 Academic Press.