摘要
Human β-defensin 3(hBD-3) is a short polypeptide with a wide range of antimicrobial activity,which was purified from human lesional psoriatic scales in 2001.To obtain high level expression in E.coli of β-defensin 3,four pairs of oligonucleotide with cosmic site were synthesised using E.coli biased codons according to the amino acid sequence of β-defensin 3, connected and amplified by PCR. The PCR product encoding human β-defensin 3 was cloned into pET30a vector.The recombinant vector was transformed into E.coli BL21(DE3)PlysS and the expression was induced by IPTG. The recombinant fusion protein was analyzed by SDS-PAGE and purified by affinity column. The mass of the fusion protein consisted of 30.9% in total bacteria proteins. The recombinant fusion protein was digested by enterokinase, resulting in the recombinant hBD-3. Antimicrobial activity analysis showed that both recombinant hBD-3 fusion protein and recombinant hBD-3 had similar potency as the native protein in suppressing growth of both gram positive bacteria S.aureus and gram negative one E.coli in a dose dependent manner.
Human β-defensin 3 （ hBD3 ） is a short polypeptide with a wide range of antimicrobial activity, which was purified from human lesional psoriatic scales in 2001. To obtain high level expression in E. coli of β- defensin 3, four pairs of oligonucleotide with cosmic site were synthesised using E. coli biased codons according to the amino acid sequence of β-defensin 3, connected and amplified by PCR. The PCR product encoding human β-defensin 3 was cloned into pET30a vector. The recombinant vector was transformed into E. coli BL21 （DE3） PlysS and the expression was induced by IPTG. The recombinant fusion protein was analyzed by SDSPAGE and purified by affinity column. The mass of the fusion protein consisted of 30.9% in total bacteria proteins. The recombinant fusion protein was digested by enterokinase, resulting in the recombinant hBD3. Antimicrobial activity analysis showed that both recombinant hBD3 fusion protein and recombinant hBD3 had similar potency as the native protein in suppressing growth of both gram positive bacteria S. aureus and gram negative one E. coli in a dose dependent manner.
出处
《中国生物化学与分子生物学报》
CAS
CSCD
北大核心
2005年第5期705-708,共4页
Chinese Journal of Biochemistry and Molecular Biology
基金
浙江省自然科学基金资助项目(No.Y204348
No.300024)~~
