title: Polycistronic expression of human platelet factor 4 with heparin-neutralizing activity in Escherichia coli.
abstract: Human platelet factor 4 (hPF4) was evaluated as a clinical alternative to protamine for heparin neutralization, a protector against radiation injury and an anti-neoplastic. To achieve high-level expression of hPF4, expression vectors pET-28a(+)-nf PF4 (n=4, 5, 6) containing n tandem repeats of PF4 were constructed and transformed into the Escherichia coli BL21(DE3) strain. A higher expression level, about 45% of the total proteins (TP), was obtained for E. coli BL21(DE3)/pET28a(+)-nf PF4 (n=4, 5, 6). The purified His-PF4 protein was further identified by cleavage with enterokinase and MS, and its heparin-neutralizing activity was determined by colony formation assay. This study represents a novel approach to large-scale production of PF4 in E. coli, one that might be applied to large-scale production of PF4 protein for possible clinical application. It also provides theoretical points for the expression and purification of other small-molecule peptides.
authors.affiliation: Department of Biochemical Engineering, School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
authors.forename: Yitao
authors.initials: Y
authors.lastname: Duan
authors.forename: Zhe
authors.initials: Z
authors.lastname: Wang
authors.forename: Wei
authors.initials: W
authors.lastname: Wu
authors.forename: Zhenjiang
authors.initials: Z
authors.lastname: Fang
authors.forename: He
authors.initials: H
authors.lastname: Huang
mesh_terms: D004765:Enteropeptidase / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D006493:Heparin / Q000378:metabolism*; D006801:Humans; D010978:Platelet Factor 4 / Q000235:genetics* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D011993:Recombinant Fusion Proteins; D010978:Platelet Factor 4; D006493:Heparin; D004765:Enteropeptidase