The cattle tick antigen, Bm95, expressed in Pichia pastoris contains short chains of N- and O-glycans

Bm95 is an antigen isolated from Boophilus microplus strains with low susceptibility to antibodies developed in cattle vaccinated with the recombinant Bm86 antigen (Gavac, HeberBiotec S.A., Cuba). It is a Bm86-like surface protein (SwissProt Accession No. P20736), which by similarity contains seven EGF-like domains and a lipid-binding GPI-anchor site at the C-terminal region. The primary structure of the recombinant (rBm95) protein expressed in Pichia pastoris was completely verified by LC/MS. The four potential glycosylation sites (Asn 122, 163, 329, and 363) are glycosylated partially with short N-glycans, from Man5GlcNAc2 to Man9GlcNAc2 of which, Man8-9GlcNAc2 were the most abundant. O-Glycopeptides are distributed mostly towards the protein N-terminus. While the first N-glycosylated site (Asn122) is located between EGF-like domains 2 and 3, where the O-glycopeptides were found, two other N-glycosylated sites (Asn329 and Asn363) are located between EGF-like domains 5 and 6, a region devoid of O-glycosylated Ser or Thr.