摘要
产肠毒素大肠杆菌(ETEC)定植于仔猪肠道的第一步是通过987P菌毛与小肠上皮细胞表面刷状缘大分子(BBV)结合。对分离的BBV进行SDS-PAGE和Ligand blot分析表明,在32～35KDa区域内有一条带能被987P菌毛探针所识别和结合,所结合的条带经胰蛋白酶消化后,通过微内径反相高效液相色谱(RP-HPLC)分离出多条主要峰带蛋白峰带,采用衬质辅助激光解吸与电离质谱法(MALDI-MS)对主要峰带进行分析,结合多肽氨基酸测序和Blast同源性比较,得到3个氨基酸基序(AETAP、ALAAAGYDVEK和LGLK),其序列与人和鼠源的组蛋白H1高度同源;来源于仔猪小肠上皮细胞BBV的H1蛋白与BBV一样都能特异性结合纯化的987P菌毛蛋白。上述结果表明,仔猪小肠上皮细胞BBV的组蛋白H1是987P菌毛蛋白的受体。
The 987P fimbriae of enterotoxigenic Escherichia coli （ETEC） mediates adhesive interactions with brush border vesicle （BBV） of the intestinal epithelial cells from the neonatal piglets. By adhering to intestinal epithelial cells, producing localized multi- plication, the 987P ETEC can progress to mucosal surface colonization and concomitant effective enterotoxin delivery. To identify the receptors for the 987P, BBV proteins from piglet intestinal villous epithelial cells were separated by SDS-PAGE and analyzed by Ligand blot, protein bands with a set of 32-35 kD recognized by the 987P fimbriae were subjected to in gel proteolysis with trypsin. The tryptic fragments were separated by microbore reversed phase HPLC（RP-HPLC）, samples shown to contain one major peak by MALDI-MS were submitted to Edman sequencing, three peptides were sequenced successfully and the all of three peptides matched the sequences of human or porcine histone H1 proteins. Porcine histone H1 proteins isolated from both piglet intestinal epithelial cells and BBV demonstrated the same SDS-PAGE migration pattern and 987P-binding properties as the 987P-specific protein receptors from piglet intestinal brush border did. The above results indicated that the 987P protein receptors are piglet BBV-derived Histone H1 proteins.
出处
《生物工程学报》
CAS
CSCD
北大核心
2008年第3期363-367,共5页
Chinese Journal of Biotechnology
基金
国家自然科学基金(No.30571374)
江苏省自然科学基金(No.BK2005049)
教育部留学回国人员科研启动基金资助项目。部分工作在美国宾夕法尼亚大学兽医学院微生物实验室完成和美国农业部USDA高新技术2002-35204-12216部分资助~~
