内生枯草芽孢杆菌SWB8 菌株β-1，3-1，4-葡聚糖酶的抗菌作用及细胞毒性

【目的】本文研究从药用植物黄姜中分离的内生枯草芽孢杆菌菌株SWB8分泌的β-1,3-1,4-葡聚糖酶的抗菌活性和细胞毒性。【方法】利用液体发酵、凝胶渗透色谱(GPC)、十二烷基-聚丙烯酰胺凝胶电泳(SDS-PAGE)和液相层析串联质谱(LC-MS/MS)等方法纯化和鉴定枯草芽孢杆菌株SWB8合成的β-1,3-1,4-葡聚糖酶;利用纸片扩散法,检测葡聚糖酶抑制临床致病性细菌和真菌生长的活性;应用MTT法和流式细胞术(FCM)评估此葡聚糖酶对人肺腺癌细胞(A549)和骨髓间质干细胞(MSCs)的细胞毒性。【结果】细菌性β-1,3-1,4-葡聚糖酶显示了广谱的抗菌活性;抗肿瘤活性主要以细胞凋亡的方式选择性的抑制人肺腺癌细胞系A549细胞的增殖,而对人骨髓间质干细胞系MSC细胞无明显影响。【结论】首次报道β-1,3-1,4-葡聚糖酶的抗菌和抗肿瘤细胞的活性。内生枯草芽孢杆菌SWB8菌株有可能成为抗菌和高效低毒的抗肿瘤药物的潜在来源。

Antibacterial effect and cytotoxicity of beta-1,3-1, 4-glucanase from endophytic Bacillus subtilis SWB8

We studied the antibiotic activity and selective cytotoxicity of β-1,3-1,4-glucanase from endophytic Bacillus subtilis SWB8. Methods ] Based on gel permeation chromatography,sodium dodecyl sulfatepolyacrylamide gel electrophoresis and liquid chromatography-tandem mass spectrometry methods,protein fragments of β-1,3-1,4-glucanase from endophytic Bacillus subtilis strain SWB8 were purified and identified.Then,β-1,3-1,4-glucanase was used to evaluate the antimicrobial activity against Staphylococcus aureus,Enterococcus faecalis,Bacillus subtilis,Escherichia coli,Salmonella typhi,Salmonella paratyphi A,Shigella dysenteriae,Candida albicans and Cryptococcus neoformans and cytotoxicity against human pulmonary adenocarcinoma cells (A549) and human bone marrow mesenchymal stem cells (MSCs) by using the disc diffusion,methyl thiazolyl tetrazolium and flow cytometry methods,respectively. Results] Bacterial β-1,3-1,4-glucanase showed broad antimicrobial spectrum against all nine bacterial and fungal strains.Furthermore,β-1,3-1,4-glucanase possessed significant anticancer activity against A549 cells that the IC50 and IC90 values were 11.5 and 20.1 μg/mL,respectively.The percentage of apoptotic A549 cells treated with different concentrations of β-1,3-1,4-glucanase was significantly increased from 4.43％ of the control to 43.1％ of 19.2 μg/mL glucanase in a dose dependent manner.In contrast,these changes could not be observed in human bone marrow mesenchymal stem cells. Conclusion ] β-1,3-1,4-glucanase could be a potential source of desirable antimicrobial agent,or anticancer compounds with higher efficiency and lower toxicity.