抗菌肽Plectasin的新型体外折叠方法与活性鉴定

A New Strategy to Activate Chemically Synthetic Plectasin in Vitro and the Identification of its Antibacterial Activity

在体外条件下,通过体外折叠体系折叠化学合成的Plectasin,经过HPLC纯化及圆二色谱仪(CD)进行结构鉴定。以耐甲氧西林金黄色葡萄球菌(MRSA),耐万古霉素肠球菌(VREF)及耐青霉素肺炎链球菌(PRSP)为实验菌株,用微量肉汤稀释法测定Plectasin对耐药菌的最小抑菌浓度(MIC)。结果:经体外折叠的Plectasin通过HPLC进行纯化,样品峰约在44.8 min出现,回收率约15%;圆二色谱鉴定显示,折叠后的Plectasin分子结构与折叠前不同,和天然构象类似;折叠活化的Plectasin对MRSA、VREF及PRSP的MIC分别为28,14,28μg/m L。通过体外折叠体系折叠化学合成的Plectasin抗菌肽,能使其获得与天然构象相当的抗菌活性。

Plectasin,the first defensin which is isolated from a fungus,has potent antimicrobial activity against gram positive pathogens,including some antibiotic-resistance strains. It is known that the Pichia pastoris expression system has been used in the expression of Plectasin. Unfortunately,expression systems are unable to guarantee a stable yield of peptides. The author＇ s objective was to activate chemically synthesized Plectasin in vitro. In their study Peletains was folded by in vitro folding system,and purified by high performance liquid chromatography（ HPLC）. And the molecular construction was identified with circular dichroism（ CD）. Antibacterial activity of folded Plectasin was tested against Methicillin-resistant Staphylococcus aureus（ MRSA） 15471118,Vancomycinresistant Enterococcus feces（ VREF） and Penicillin-resistant Streptococcus pneumonia（ PRSP） 31355. The folded Plectasin was purified through HPLC at 44. 8 min with recovery rate of approximately 15%. CD has shown that the molecular structure of chemically synthesized Plectasin was changed from its unfolded counterpart and was consistent with its natural form. The antimicrobial activities of folded Plectasin against MRSA,VREF and PRSP were 28,14,28 μg/m L respectively,which were similar to those of inducible Plectasin in Pichia pastoris as compared with their previous research. In conclusion,chemically synthesized Plectasin was successfully folded by the folding system in vitro,giving it as much antimicrobial activity as its natural form.