Determination of glycosylation sites using a protein sequencer and deglycosylation of native yeast invertase by endo-beta-N-acetylglucosaminidase.
Endo-beta-N-acetylglucosaminidase from Arthrobacter protophormiae was tested for its capacity to release N-linked sugar chains from native yeast invertase. The enzyme liberated about 80% of the sugar chains from the native invertase. Deglycosylated invertase was digested by chymotrypsin or pepsin, and twelve N-acetylglucosamine-containing glycopeptides were isolated. The amino acid sequences of these glycopeptides were analyzed by a protein sequencer, and the elution position of 4-L-aspartylglycosylamine was directly identified by conventional sequencing. The endo-beta-N-acetylglucosaminidase was found to remove mainly nine sugar chains from native invertase.
Takegawa
K
K
Department of Bioresource Science, Faculty of Agriculture, Kagawa University, Japan.
Yoshikawa
M
M
Mishima
T
T
Nakoshi
M
M
Iwahara
S
S
D000595:Amino Acid Sequence; D002240:Carbohydrate Sequence; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006031:Glycosylation; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000378:metabolism*; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D043324:beta-Fructofuranosidase
D006026:Glycoside Hydrolases; D043324:beta-Fructofuranosidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase