title: Purification and modification by polyethylene glycol of a new human basic fibroblast growth factor mutant-hbFGF(Ser25,87,92).
abstract: The mutant of human basic fibroblast growth factor (hbFGF), hbFGF(Ser25,87,92), which was constructed by replacing the cysteine residues at the positions of the 25th, the 87th and the 92nd with serine residues, was coupled to polyethylene glycol (PEG) with a molecular size of 20 kDa (20K) (PEG(20K)) to obtain hbFGF derivative, PEG(20K)-hbFGF(Ser25,87,92). The optimal modified reaction was conducted at 12 degrees C for 12h with the molar ratio of PEG(20K) to hbFGF(Ser25,87,92) of 30:1. The result of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) showed that the modification rate was up to 60%. The PEGylated product retained binding affinity to heparin and could be purified by heparin affinity chromatography. Compared to hbFGF mutant, purified PEG(20K)-hbFGF(Ser25,87,92) retained about 34% of mitogenic activity. Heat-stability assay indicated that the modified product was more stable than the native protein at the temperature of 37 degrees C.
authors.affiliation: Chinese-American Research Institute for Diabetes Complications, School of Pharmacy, Wenzhou Medical College, Wenzhou 325035, China.
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Wu
authors.forename: Xiaoju
authors.initials: X
authors.lastname: Liu
authors.forename: Yecheng
authors.initials: Y
authors.lastname: Xiao
authors.forename: Zhifeng
authors.initials: Z
authors.lastname: Huang
authors.forename: Jian
authors.initials: J
authors.lastname: Xiao
authors.forename: Shaoqiang
authors.initials: S
authors.lastname: Lin
authors.forename: Lu
authors.initials: L
authors.lastname: Cai
authors.forename: Wenke
authors.initials: W
authors.lastname: Feng
authors.forename: Xiaokun
authors.initials: X
authors.lastname: Li
mesh_terms: D004591:Electrophoresis, Polyacrylamide Gel; D016222:Fibroblast Growth Factor 2 / Q000235:genetics / Q000302:isolation & purification* / Q000494:pharmacology; D006801:Humans; D008938:Mitosis / Q000187:drug effects; D016297:Mutagenesis, Site-Directed; D011092:Polyethylene Glycols / Q000737:chemistry*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology
chemical_list: D011994:Recombinant Proteins; D016222:Fibroblast Growth Factor 2; D011092:Polyethylene Glycols