title: [Pichia pastoris X-33 with OCH1 gene deletion and its expression of glycoprotein GM-CSF].
abstract: OBJECTIVE
  Glycoproteins derived from yeast expression systems are usually hyperglycosylated and contain non human N-glycans of the high mannose type. alpha-1, 6-mannosyltransferase (och1p) plays a key role in modifying glycoproteins with high mannose-type N-glycans. Therefore, we engineered a P. pastoris X-33 strain with OCH1 gene deletion. This strain was further used as a host for production of glycoproteins with smaller N-glycans.
  
  
  METHODS
  First, we knockout the URA3 gene of P. pastoris X-33 which encoding orotidine-5'-phosphate decarboxylase by double homologous recombination. Then, we knockout OCH1 gene using URA3 gene as a selecting marker, and obtained X-33 (och1-) strain. After that, this mutant strain was used to expression the glycoprotein granulocyte-macrophage colony-stimulating factor (GM-CSF).
  
  
  RESULTS
  Different from hyperglycosylated GM-CSF expressed in wild type P. pastori X-33, the glycoprotein expressed in X-33 (och1-), containing smaller N-glycan.
  
  
  CONCLUSIONS
  The results suggested that X-33 (och1-) strain can be used as an expression host for production of glycoproteins lacking the outer-chain hypermannoses, and a host could be used for further N-glycosylation engineering.
authors.affiliation: School of Biotechnology, Jiangnan University, Wuxi 214122, China. zhangdachenghb@163.com
authors.forename: Dacheng
authors.initials: D
authors.lastname: Zhang
authors.forename: Yongli
authors.initials: Y
authors.lastname: Xu
authors.forename: Xin
authors.initials: X
authors.lastname: Xin
authors.forename: Ruiyu
authors.initials: R
authors.lastname: Zhu
authors.forename: Jian
authors.initials: J
authors.lastname: Jin
mesh_terms: D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal; D055786:Gene Knockout Techniques; D006023:Glycoproteins / Q000096:biosynthesis / Q000235:genetics; D006031:Glycosylation; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000096:biosynthesis / Q000235:genetics*; D008358:Mannose / Q000378:metabolism; D008364:Mannosyltransferases / Q000235:genetics* / Q000378:metabolism; D009964:Orotidine-5'-Phosphate Decarboxylase / Q000235:genetics; D010843:Pichia / Q000235:genetics* / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D005656:Fungal Proteins; D006023:Glycoproteins; D011134:Polysaccharides; D011993:Recombinant Fusion Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D008364:Mannosyltransferases; C058478:alpha 1,6-mannosyltransferase; D009964:Orotidine-5'-Phosphate Decarboxylase; D008358:Mannose