title: PEGylation enhancement of pH stability of uricase via inhibitive tetramer dissociation.
abstract: OBJECTIVE
  Previously, PEGylated uricase was demonstrated to maintain catalytic activity at pH 5.8, the isoelectric point of uricase, where native uricase ceases to function. To find out whether PEGylation could enhance pH stability of uricase, the enzyme activity to pH curve was completely characterized.
  
  
  METHODS
  Complete characterization of the enzyme activity to pH curve, indicating an inverted bell-shaped relationship not previously documented, is presented. PEGylation enhancement of uricase stability at a pH lower than that commonly found in the liver, can be explored by dynamic dissociation of uricase using ultrafiltration and size-exclusion chromatography.
  
  
  RESULTS
  The results suggest the role of PEGylation in enhanced pH stability is via inhibition of subunit disintegration. The mechanism of this effect is characterized by the wrapping of PEG chains around uricase, providing a flexible shell preventing subunit disintegration. The presence of notable PEGylation-induced changes in uricase supports this mechanism and include improved enzyme-substrate affinity and elevated thermal stability.
  
  
  CONCLUSIONS
  Characterization of PEGylated uricase provides a basis for the rational design of therapeutic PEGylated proteins.
authors.affiliation: College of Life Science and Technology, China Pharmaceutical University, Nanjing, China.
authors.forename: Hong
authors.initials: H
authors.lastname: Tian
authors.forename: Yuan
authors.initials: Y
authors.lastname: Guo
authors.forename: Xiangdong
authors.initials: X
authors.lastname: Gao
authors.forename: Wenbing
authors.initials: W
authors.lastname: Yao
mesh_terms: D002175:Candida / Q000201:enzymology; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004058:Diffusion; D004355:Drug Stability; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000032:analysis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006074:Gout Suppressants / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D006358:Hot Temperature / Q000009:adverse effects; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D011092:Polyethylene Glycols / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000032:analysis / Q000378:metabolism; D012995:Solubility; D014462:Ultrafiltration; D014503:Urate Oxidase / Q000032:analysis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D004800:Enzymes, Immobilized; D005656:Fungal Proteins; D006074:Gout Suppressants; D011994:Recombinant Proteins; C469709:rasburicase; D011092:Polyethylene Glycols; D014503:Urate Oxidase; C031545:Pegloticase