title: Determination of N-glycosylation site and glycan structures of pectin methylesterase in jelly fig (Ficus awkeotsang) Achenes.
abstract: Pectin methylesterase (PME) in jelly fig (Ficus awkeotsang) achenes is an N-glycosylated enzyme responsible for the gelation of jelly curd. A recombinant jelly fig PME was overexpressed in Escherichia coli and confirmed by immunodetection and LC-nanoESI-MS/MS analysis. To identify the N-glycosylation site, native PME and its deglycosylated and recombinant forms, which lacked glycan, were purified and subjected to comparative MALDI-MS mapping of the corresponding tryptic fragments. The results showed that N-glycosylation occurred at Asn(153) of the mature jelly fig PME, the only glycosylation site predicted by its sequence analysis. The major N-glycans released from the native PME by PNGase F were identified by MS/MS analyses as xylosylated, noncore fucosylated pauci-mannose, and complex-type structures. Molecular modeling of the 3D structure of jelly fig PME indicated that the N-glycan was putatively attached to the back region of the active site of this enzyme.
authors: Graduate Institute of Biotechnology, National Chung-Hsing University, Taichung, Taiwan.
authors: Eric S L
authors: ES
authors: Hsiao
authors: Jeff C F
authors: JC
authors: Chen
authors: Hsien-Yu
authors: HY
authors: Tsai
authors: Kay-Hooi
authors: KH
authors: Khoo
authors: Shui-Tein
authors: ST
authors: Chen
authors: Jason T C
authors: JT
authors: Tzen
mesh_terms: D000595:Amino Acid Sequence; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D030681:Ficus / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D006031:Glycosylation; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011134:Polysaccharides / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D010940:Plant Proteins; D011134:Polysaccharides; D002265:Carboxylic Ester Hydrolases; C022323:pectinesterase