COVALENT STRUCTURE OF BOVINE BRAIN CALRETICULIN

The covalent structure of bovine brain calreticulin, a major Ca2+- bin ding protein in the lumen of the endoplasmic reticulum, was determined by analysis of the purified protein. The protein consisted of 400 ami no acids, with an N-linked oligosaccharide attached to the polypeptide chain. The polypeptide sequence determined was compatible with the se quence of calreticulin deduced from cDNA of different sources, with a number of differences presumably due to species-specific amino acid su bstitutions. The protein retained the C-terminal tetrapeptide, KDEL, i nvolved in retention of proteins resident in the endoplasmic reticulum , whereas the N-terminal signal peptide predicted from the cDNA sequen ce had been removed in the purified protein. The bovine brain protein contained a high-mannose type of oligosaccharide attached to Asn(162), which is typical of resident endoplasmic reticulum proteins. The carb ohydrate moiety was heterogeneous and had the composition GlcNAc(2)Man (4-9), of which GlcNAc(2)Man(5) was the most abundant in the bovine br ain preparation. Glycosylation of calreticulin, however, appeared to b e a species-specific modification, as Asn(162) is replaced by Asp in t he sequences already determined for a number of species. Analysis of t he purified protein also identified an intramolecular disulphide bridg e between Cys(120) and Cys(146).