壳聚糖诱导下浅玫瑰色链霉菌菌体蛋白差异表达分析

Analysis of two-dimensional electrophoresis for total proteins from Streptomyces roseolus by induction of chitosan

建立了有效分析浅玫瑰色链霉菌菌体全蛋白质的方法,使用分离范围为p H 4-7的IPG胶条、2D clean-up试剂盒法纯化方法、80μg蛋白质上样量和银染的双向电泳技术,获得了低背景、高分辨率和重复率、无明显条纹的浅玫瑰色链霉菌全蛋白质双向电泳图谱。通过Image Master2D Platinum7.0软件对壳聚糖诱导条件下浅玫瑰色链霉菌菌体总蛋白质的电泳图谱进行匹配分析,在诱导菌株图谱上有723±14个清晰可见的蛋白质点可供差异分析。与未诱导菌株的双向电泳图谱对比结果显示：18个蛋白质点在诱导菌株中差异表达,其中14个蛋白质点的表达量上调,4个蛋白质点的表达量下调,选取10个差异表达量在5倍以上的蛋白质点进行质谱鉴定,结果显示,延长因子Tu、RNA聚合酶、分子伴侣等蛋白在壳聚糖诱导下表达量增加,表明它们在浅玫瑰色链霉菌代谢壳聚糖的过程中起重要作用。

Two-dimensional gel electrophoresis method for analysis of total proteins from Streptomyces roseolus was established in this paper as follows： IPG strips with p H 4-7,2D clean-up kit purification protocol,80μg protein sample volume and silver staining.2D-gels were obtained with low background,high resolution,good reproducibility and without obvious stripes.The 2-DE maps of total proteins from S.roseolus by induction of chitosan were analyzed by Image Master2 D Platinum7.0,which showed 723 ± 14 clearly visible protein spots.Compared with 2-DE maps of total proteins from S.roseolus incubated with no induction,18 proteins detected in induced strain were in differential expression.Among these,14 proteins were upregulated and 4 proteins were down-regulated.10 proteins whose expression levels differed more than 5 times were selected and identified by mass spectrometry.The results showed that the expressions of proteins such as elongation factor Tu,RNA polymerase,chaperone and so on were increased when incubating with chitosan,which suggested that they may be involved in the process of chitosan metabolism in S.roseolus.