Prokaryotic Expression, Purification and Biological Activity of Human β-NGF

Objective To express human β-nerve growth factor(β-NGF)in E. coli, purify the expressed product and deter-mine its biological activity. Methods β-NGF gene fragment was amplified by PCR using human peripheral blood genomic DNA as a template, then cloned, sequenced and inserted into expression vector pET-28a(+). The constructed recombinant plasmid pET-28a-β-NGF was transformed to E. coli BL21(DE3)for expression under induction of IPTG. The expressed product was purified by molecular sieve chromatography, then re-naturalized and determined for biological activity by chick embryo dorsal root ganglion culture test. Results Restriction analysis and sequencing proved that the gene fragment encoding human β-NGF mature peptide was amplified by PCR. The expressed product mainly existed in a form of inclusion body, contained about 23. 5% of total somatic protein, reached a purity of more than 90% after purification and promoted the growth of tubercle of chick embryo dorsal root ganglion significantly. The expression level of target protein was 4. 8 mg / L bacterial liquid. Conclusion Human β-NGF was successfully expressed in E. coli and showed high biological activity after purification.