家蚕幼虫5龄期中肠蛋白质组学

Proteomics of Midgut of Fifth-instar Silkworm Larva

利用双向电泳技术对家蚕幼虫5龄期第2天、第5天和第7天的中肠蛋白质进行分离,并利用ImageMaster 2D Platinum对所分离得到的蛋白图谱进行比较,并对一些蛋白质斑点进行了质谱鉴定。研究发现,家蚕中肠蛋白质具有区别于家蚕其他组织的明显特征:蛋白质大多分布在PI值4-8、分子量20～70 kD的区域,且分布不均匀,主要集中在酸性一侧,这一特点在家蚕5龄期第7天的图谱尤为明显。5龄期家蚕第2天的蛋白质斑点数目为869个,而到第5天增加到966个,新增蛋白数目97个,进一步观察发现增加的蛋白主要分布在PI值6-9,分子量20～40kD区间;随着进入幼虫成熟期,蛋白质斑点数目明显减少,第7天斑点数仅为420个,比第5天减少了56.5%。这些结果说明家蚕中肠组织蛋白质组成在5龄早、中、晚期经历了较大变化,暗示这可能与中肠的功能相适应。从MALDI-TOF-MS鉴定的斑点发现了构成家蚕中肠组织的一些新的部分结构蛋白和一些可能与消化、吸收相关的蛋白,还发现一些能够抵御外界微生物入侵的相关蛋白。这些结果为进一步认识家蚕中肠提供了重要的理论基础。

We used high resolution two-dimensional polyacrylamide gel electrophoresis and ImageMaster 2D platinum techniques to study the total midgut proteins of silkworm, Bombyx mori in the 2nd, 5th and 7th day of the fifth instar stage. The characteristics of midgut proteomic differed from other reported silkworm tissues. Most protein spots were unevenly distributed in the area from 20 to 70 kD and PI between 4 and 8. Among them most spots located in acidic area, and this characteristic was observed in the map of midgut in the 7th day. There were 869 spots in the 2D map of midgut in the 2nd day, and 966 in 5th day. During this period, 97 new spots were generated, and mainly located in the area of pI 6-9 with molecular size between 20 and 40 kD. Along with larval maturation, the spots in the map decreased obviously, with 420 in the 7th day, 56.5% less than that of the 5th day. These results indicated that the proteins in the midgut had big change at the early, middle and late 5th stage, implying their adaptation in the larval midgut. Some spots were excised and identified by using MALDI-TOF MS, and they were involved in larval body structure, metabolism and immunity.