GLYCOSYLATION OF HUMAN ALPHA(1)-ANTITRYPSIN IN SACCHAROMYCES-CEREVISIAE AND METHYLOTROPHIC YEASTS

Human alpha(1)-antitrypsin (alpha(1)-AT) is a major serine protease in hibitor in plasma, secreted as a glycoprotein with a complex type of c arbohydrate at three asparagine residues. To study glycosylation of he terologous proteins in yeast, we investigated the glycosylation patter n of the human alpha(1)-AT secreted in the baker's yeast Saccharomyces cerevisiae and in the methylotrophic yeasts, Hansenula polymorpha and Pichia pastoris. The partial digestion of the recombinant alpha(1)-AT with endoglycosidase H and the expression in the mnn9 deletion mutant of S. cerevisiae showed that the recombinant alpha(1)-AT secreted in S. cerevisiae was heterogeneous, consisting of molecules containing co re carbohydrates on either two or all three asparagine residues. Besid es the core carbohydrates, variable numbers of mannose outer chains we re also added to some of the secreted alpha(1)-AT. The human alpha(1)- AT secreted in both methylotrophic yeasts was also heterogeneous and h ypermannosylated as observed in S. cerevisiae, although the overall le ngth of mannose outer chains of alpha(1)-AT in the methylotrophic yeas ts appeared to be relatively shorter than those of alpha(1)-AT in S. c erevisiae. The alpha(1)-AT secreted from both methylotrophic yeasts re tained its biological activity as an elastase inhibitor comparable to that of alpha(1)-AT from S. cerevisiae, suggesting that the different glycosylation profile does not affect the in vitro activity of the pro tein. (C) 1998 John Wiley & Sons, Ltd.