Mass spectrometric characterization and glycosylation profile of bovine pancreatic bile salt-activated lipase.

Abstract

We developed a procedure for the large scale isolation of bovine bile salt-activated lipase (BAL) for its crystallization [Wang, X., et al. (1997) Structure 5, 1209-1218] and also carried out a study on the molecule's glycosylation profile for a better deduction of the structure of the enzyme. Mass spectrometric analysis of the CNBr-generated peptides indicated that only one (Asn-361) of the two potential N-glycosylation sites (Asn-187 and Asn-361) with NXT motif is glycosylated. The analysis of the isolated CNBr peptide containing Asn-361 showed that it existed in three glycoforms in a ratio of 1.0:2.8:1.0, with oligosaccharide moieties weighing 1900.1, 2045.2, and 2336.4 Da, respectively. The major oligosaccharide chain contained mannose: galactose:N-acetylglucosamine:fucose:sialic acid in a molar ratio of 2:2:4:2:1. It was also determined that the potential O-glycosylated peptide (CB13) is not O-glycosylated and, in addition, it was found that there was microheterogeneity in the C-terminus of the isolated bovine BAL. The results obtained from this mass spectrometric study combined with the X-ray crystallographic studies provide more precise structural information on BAL. The procedure described here for the mass spectrometric analysis of CNBr-generated peptides also has general applicability for analysis of the glycosylation profile of glycoproteins and the C-terminal peptide structure of proteins.

Cite this paper

@article{Wang1998MassSC, title={Mass spectrometric characterization and glycosylation profile of bovine pancreatic bile salt-activated lipase.}, author={Changjun Wang and Kenneth W. Jackson and Ali Dashti and Deborah Downs and X Zhang and Jing Jin Tang}, journal={Protein expression and purification}, year={1998}, volume={12 2}, pages={259-68} }