No abstract

Complementary DNA encoding hen egg white lysozyme (HEWL) was subjected to site-directed mutagenesis to introduce two N-linked glycosylation sites (Asn19-Try20-Thr21 and Asn49-Ser50-Thr51) into both positions 19 and 49 by substituting Arg-21 with Thr and Gly-49 with Asn, respectively. The double-glycosylated lysozyme R21T/G49N was expressed in Saccharomyces cerevisiae carrying the yeast expression plasmid inserted the double-mutant HEWL cDNA. The mutant lysozyme mainly secreted a polymannosyl form with a small amount of two oligomannosyl forms. The polymannosyl lysozyme R21T/G49N was glycosylated at two positions, 19 and 49, with a polymannosyl and an oligomannosyl chain. The lengths of the polymannosyl and oligomannosyl chains attached to R21T/G49N were approximately 272 and 18 mannose residues, respectively. The R21T/G49N showed better emulsifying properties than two types of single-polymannosyl lysozymes R21T and G49N. With regard to single-polymannosyl lysozyme, G49N showed somewhat better emulsifying properties than R21T. In addition, the cleavage of polymannosyl chain from lysoyme with endo-β-N-acetylglucosaminidase resulted in a dramatic decrease in the emulsifying properties of polymannosyl lysozymes. Keywords: Lysozyme; yeast expression system; genetic modification; polymannosylation; double glycosylation; emulsifying properties

No abstract

No abstract