Expression Of Antibacterial Peptide SMAP-29 By SUMO Technology In E.coliand Analysis Of Its Antibacterial Activity

Antibacterial peptides is encoded by the biological specific gene which is a kind of small molecular peptides. It has the function of the violation against outside microorganisms and it is an important component part of the innate immune system. Because of its good heat stability, strong antibacterial activity, broad-spectrum antibacterial and antimicrobial resistance, it had been widely attention.Antibacterial peptide SMAP-29 derived from sheep myeloid cells of Cathelicidins precursor protein C interceptor 29 amino acid residues by Bagella and Margherita Zanetti. It belongs to the class Cathelicidin antimicrobial peptides with a-helical structure domain, which has a strong resistance to bacteria, fungi, virus, and other biological function.According to SMAP-29 amino acid sequence and E.coli codon preferences, we designed the gene of SMAP-29, constructed recombinant expression vector pSUMO-SMAP-29; optimized expression induced conditions of fusion protein SUMO-SMAP-29; expressed and purified fusion protein SUMO-SMAP-29 in E.coli BL21(DE3); also expressed and purified SUMO protease 1 in E.coli BL21(DE3); separated and purified antibacterial peptide SMAP-29 by Ni-NTA; determined the antimicrobial activity of SMAP-29 in vitro.The main results were as follows:(1) We constructed recombinant expression vector pSUMO-SMAP-29 successfully.(2) We found that IPTG 0.5mM, 30 ℃ and 3h was the optimal induction condition.(3) We obtained 16 mg SUMO-SMAP-29 per liter of culture medium through the Ni column affinity chromatography.(4) We obtained 115 mg SUMO protease1 per liter of culture medium through the Ni column affinity chromatography.(5) Kirby-Bauer test and the MIC experiments showed that recombinant antimicrobial peptide SMAP-29 displayed antibacterial activity. The MIC is 32 ug/mL for E.coli ATCC25922. But the antibacterial activity is not obvious for S.a ureus ATCC25923.