title: Characterization of an extracellularly derived α-mannosidase from the liquid exudate of the sclerotia of Sclerotinia sclerotiorum (Lib.) de Bary.
abstract: Class I α-mannosidases play an important role in co- and post-translational N-glycosylation modification of proteins, and also in glycoprotein glycan hydrolysis. Herein, we investigated a protein named Man-41, from liquid exudate droplets secreted on the surface of developing sclerotia by Sclerotinia sclerotiorum. The protein was identified by MALDI-TOF mass spectrometry to be a α-mannosidase. The full-length open reading frame of Man-41 consists of 1581 bp, encoding 526 amino acid residues and containing a putative signal peptide at amino acid residues 1-20, and a conserved sequence at residues 50-521. Man-41 was classified into glycoside hydrolase family 47 (GH47) by clustering analysis. The catalytic residues include Glu(125), Arg(129), Asp(270), Ser(271), Glu(274), Arg(420), Glu(422), Glu(425), Glu(485), Thr(514), and Glu(515), which are conserved in all Class I α-1,2-mannosidases. Recombinant Man-41 protein had 26.67 ± 2.18 U/mg of α-mannosidase activity, about one-half of intracellular mannosidase activity of sclerotia. In conclusion, this is the first time that mannosidase has been identified in an extracellular fluid and Man-41 is also a new member of GH47 with Ca(2+)-dependent characteristics. This work lays the foundation for further study of the function of Man-41 in sclerotial development.
authors.affiliation: Hunan Provincial Key Laboratory of Crop Germplasm Innovation and Utilization, Hunan Agricultural University, Changsha, 410128, Hunan, China.
authors.forename: Zhengli
authors.initials: Z
authors.lastname: Liu
authors.forename: Ran
authors.initials: R
authors.lastname: Wei
authors.forename: Wen
authors.initials: W
authors.lastname: He
authors.forename: Ying
authors.initials: Y
authors.lastname: Ruan
authors.forename: Chunlin
authors.initials: C
authors.lastname: Liu
mesh_terms: D000595:Amino Acid Sequence; D001203:Ascomycota / Q000201:enzymology*; D020134:Catalytic Domain; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D043323:alpha-Mannosidase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D043323:alpha-Mannosidase