Polycistronic Expression of Human Platelet Factor 4 with Heparin-Neutralizing Activity in Escherichia coli

• Published in: Bioscience, biotechnology, and biochemistry Vol. 76; no. 10; pp. 1855 - 1860
• Main Authors: DUAN, Yitao, WANG, Zhe, WU, Wei, FANG, Zhenjiang, HUANG, He
• Format: Journal Article
• Language: English
• Published: Tokyo Japan Society for Bioscience, Biotechnology, and Agrochemistry 2012; Japan Society for Bioscience Biotechnology and Agrochemistry; Oxford University Press
• Subjects: anti-neoplastic; Biological and medical sciences; biotechnology; chemokine CXCL4; Colonies; enteropeptidase; Enteropeptidase - metabolism; Escherichia coli; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; Gene Expression; Genetic Engineering - methods; Genetic Vectors - genetics; heparin; Heparin - metabolism; Humans; neutralization; neutralizing heparin; platelet factor 4; Platelet Factor 4 - genetics; Platelet Factor 4 - metabolism; radiation injury; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; small pharmaceutical proteins; tandem expression; Human; Drug; tandem expression; Escherichia coli; neutralizing heparin; Malignant tumor; Protein; anti-neoplastic; Platelet; platelet factor 4; Bacteria; Heparin; small pharmaceutical proteins; Cancer; Enterobacteriaceae
• ISSN: 0916-8451; 1347-6947; 1347-6947
• DOI: 10.1271/bbb.120267

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.