新疆家蚕抗菌肽Cecropin-XJ与细菌DNA相互作用的光谱研究

抗菌肽的抗菌机理研究主要集中在抗菌肽与细菌细胞膜作用方面,抗菌肽是否与细菌的染色体DNA作用尚不清楚。为了探讨新疆家蚕抗菌肽Cecropin-XJ抗细菌的作用机理,利用紫外光谱及以溴化乙锭(EthidiumBromide,EB)为荧光探针的荧光光谱方法研究抗菌肽Cecropin-XJ与金黄色葡萄球菌DNA在体外的相互作用,计算获得抗菌肽与DNA的结合常数和成键位点数。结果显示,抗菌肽使DNA发生了明显的增色效应,并使DNA的荧光强度增强,抗菌肽能与EB竞争性的结合DNA,表明抗菌肽可能与DNA双螺旋的沟槽结合;在抗菌肽存在下,DNA与EB作用的结合常数和成键位点数都发生变化,表明抗菌肽以嵌入和非嵌入两种方式与DNA相互作用。文章从分子水平上初步阐述了抗菌肽与细菌DNA的作用模式和结构特征,为深入研究抗菌肽的作用机理奠定了基础。

Study on the Interaction Mechanism of Antimicrobial Peptide Cecropin-XJ in Xinjiang Silkworm and Staphylococcus Aureus DNA by Spectra

Nowadays many of antimicrobial peptides have been extensively studied in order to elucidate their antimicrobial mode of action.Much of the research focused on mechanisms of cytoplasmic membrane disruption has been proposed for antimicrobial peptides,but it is not known whether their antimicrobial mode of action is due to their effects on bacterial chromosome.To obtain more information about the possible mechanisms,Cecropin-XJ,a kind of antimicrobial peptide from Xinjiang silkworm,was used as subject and prepared by purification of Pichia yeast fermentation containing cecropin-XJ gene expression product.Subsequently,ultraviolet absorption spectra were employed to investigate whether this antibacterial function is due to Cecropin-XJ works on S.aureus DNA in vitro.The increase in absorbance of DNA samples at 260nm due to the addition of Cecropin-XJ was measured.It is called hyperchromicity of DNA,which can provide a direct measure of the degree of base-pair unstacking.The unstacking results in the loss of duplex helix,and then leads to the duplex helix becoming relaxing.At the same time,the interaction mode was studied by using ethidium bromide(EB) as an extrinsic fluorescence probe.With the addition of Cecropin-XJ,the intensity of intrinsic fluorescence absorbance of DNA at 307 nm increases greatly.It is suggested that the addition of Cecropin-XJ may unstack base-paire of DNA exposing the fluorescent amino acide,leading to the enhancing of DNA fluorescence intensity.Subsequently the competition between Cecropin-XJ and EB to combine with DNA was found.It is suggested that the style was groove binding and intercalation in the interaction of Cecropin-XJ and double helix DNA.Furthermore,in this study,the binding constant and binding number of Cecropin-XJ complex with DNA were determined.There were different binding constant and number of EB complex with DNA because of the the addition of Cecropin-XJ.It is showed that the interaction of Cecropin-XJ and DNA was based on intercalation or non-intercalation.Meanwhile,these results help explain the molecule mechanism of antimicrobial peptide from the interaction style and structure characteristic of Cecropin-XJ and S.aureus DNA.These findings could contribute to further investigation on the mechanism of action of Cecropin-XJ.