Hydrothermal vent tube worm is the one of the most typical hydrothermal species. In previous study,we constructed cDNA library of tube worm(Ridgeia piscesae) and sequencing result showed that it contains 23 chitin-binding proteins( CBPs). With high transcription level and diversity,each protein contains 1 ~3 different types of chitin binding domain ( s). In view of the important role in chitin degradation and the special habitats of the tube worms , we further studied the functional characterization of 4 typical chitin-binding proteins. After analyzed, the sequences of signal peptides were cut off and the fragments encoding chitin-binding proteins were amplified by PCR, inserted into pIZ-FLAG Vector and successfully expressed in Hi5 cells. In the chitin affinity assay,the results demonstrated that four FLAG-CBPs can bind to a-chitin respectively. Meanwhile, the supernatant of cell lysate including the expressed product FLAG-CBPs was able to enhance the activity of chitinase in the process of degrading a-chitin and β-chitin respectively. All the results suggest that four CBPs may act as a-chitin binding proteins and may enhance the activity of chitinase. In-depth study will be taken to verify other biological functions of chitin binding proteins and optimize their ability of catalyzation.%本文应用PCR技术对四种管状蠕虫几丁质结合蛋白进行了扩增,进而连接到pIZ-FLAG构建了重组表达载体,并在昆虫细胞中实现了几丁质结合蛋白的异源重组表达.我们对几丁质结合蛋白的功能进行了初步研究,亲和活性实验结果表明这四种几丁质结合蛋白对α-chitin具有亲和活性,并且可以辅助几丁质酶,对降解α-chitin和β-chitin均有不同程度的促进作用,且作用效果明显.结果表明,这四种几丁质结合蛋白均为α型,且可以促进几丁质酶的活性,对降解几丁质多糖具有巨大的应用潜力和应用价值.
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