甘蔗β-1,3-葡聚糖酶基因的电子克隆与分析

In Silico Cloning and Bioinformatic Analysis of β-1,3-glucanase Gene(ScBG) from sugarcane(Saccharum spp.)

以高粱β-1,3-葡聚糖酶基因(β-1,3-glucanase gene)cDNA序列为探针,搜索甘蔗EST数据库,而后通过电子克隆技术,拼接获得甘蔗β-1,3-葡聚糖酶基因ScBG。采用生物信息学方法,对该基因编码蛋白从氨基酸组成、理化性质、跨膜结构域、卷曲螺旋、亚细胞定位、信号肽、功能域及高级结构等方面进行了预测和分析。结果表明:ScBG基因全长1270bp,包含一个长达1011bp的完整开放读码框(open reading frame,ORF),编码336个氨基酸,分子量为34.8KD,理论等电点为4.98。该蛋白质很可能是胞外定位的诱导物释放型酸性葡聚糖酶,是一种稳定的分泌蛋白,且可信度达最高等级1。该蛋白属于糖苷水解酶第17家族,含有N端信号肽,在第7～29位氨基酸处含有跨膜信号区,在第31～321位氨基酸处含有糖苷水解酶17家族结构域,含2个主要的功能结构域。10个物种ScBG蛋白氨基酸序列的同源性分析表明,甘蔗ScBG基因编码蛋白与高粱β-1,3-葡聚糖酶基因的编码蛋白的同源性最高,达79.82%。以上研究结果为ScBG基因下一步的分子克隆、功能鉴定和应用提供基础。

Abstract：Based on in silico cloning, a novel sugarcane β-1,3 -glucanase gene, termed as ScBG, was identified by using β- 1,3 - glucanase cDNA sequence of Sorghum bicolor as a probe to search the sugarcane ESTs data- base. Several characters of ScBG protein, including the composition of amino acid sequence, physical and chemical properties, transmembrane domain, coiled coil, subcellular localization, signal peptide, functional domain, and secondary and tertiary structure were predicted and analyzed by bioinformatics tools. The results showed that this ScBG gene, which was 1 270 bp long, contained a complete ORF of I 011 bp. It encoded the 336 amino acid poly- peptide with a molecular weight of 38.4 KD and the isoelectric point of 4.98. Besides, the protein was a stable se- Cretory protein and was most likely to be an inducer - released extracellular acid glucanase, credibility to the high- est level 1. ScBG belonged to glycoside hydrolase family 17 and contained a N - terminal signal peptide and 2 main function domain. At the loci of 7 -29 amino acids, ScBG contained transmembrane signal area while at the loci of 31 -321, it contained glycoside hydrolase structure domain specific in family 17. When comparing ScBG homolo- gous amino acid sequences among sugarcane and other plant species, the result showed that ScBG protein had the highest similarity with that of Sorghum bicolor, which was 79.82%. This study aims to provide the basis for the molecular cloning, functional analysis and application of this gene in the future.