ISOLATION AND EXPRESSION OF MURINE AND RABBIT CDNAS ENCODING AN ALPHA-1,2-MANNOSIDASE INVOLVED IN THE PROCESSING OF ASPARAGINE-LINKED OLIGOSACCHARIDES

We have isolated a full-length cDNA clone encoding a murine alpha1,2-m annosidase involved in the processing of mammalian Asn-linked oligosac charides. Oligonucleotide primers were designed based on peptide seque nces derived from the purified rabbit liver enzyme and were used to ge nerate a 1011-base pair probe using the polymerase chain reaction. Thi s probe was used to isolate clones from rabbit and mouse cDNA librarie s. The full-length murine cDNA clone encodes a 655-amino acid type II transmembrane protein with a 43-amino acid cytoplasmic tail, a single transmembrane domain, and a large COOH-terminal catalytic domain conta ining two potential N-glycosylation sites. Stable transfection of the murine alpha1,2-mannosidase cDNA into mouse L cells resulted in a appr oximately 22-fold overexpression of alpha1,2-mannosidase activity. Thr ee transcripts were detected in rabbit tissues, whereas two were found in rat and mouse tissues. The sequences of the rabbit and mouse cDNA clones indicate that the multiple transcripts differ in the length of their 3' sequences as a result of the use of multiple polyadenylation signals. Immunolocalization detected cross-reactive material in a juxt anuclear pattern consistent with the Golgi complex. The catalytic port ion of the murine alpha1,2-mannosidase was found to bear a strong simi larity to the processing alpha1,2-mannosidase from Saccharomyces cerev isiae.