灵芝漆酶注释基因Lacc1的生物信息学分析及其Cu^(2+)诱导表达

Bioinformatics analysis of putative laccase gene Lacc1 from Ganoderma lucidum and Cu^(2+)induced gene expression

为验证Lacc1基因的功能,揭示灵芝漆酶基因及其启动子结构与其转录表达的内在关系,预测了Lacc1的氨基酸序列结构及灵芝漆酶注释基因Lacc1的上游启动子区域,研究已测序的菌株——灵芝P9在Cu2+诱导条件下漆酶基因Lacc1的表达情况。Protein Blast分析结果表明,Lacc1含有3个铜氧化酶(Cu-oxidase)保守结构域,与Polyporus ciliatus漆酶lcc3-3的相似性最高,为71%,并具有高度保守的漆酶特征序列L1-L4;亚细胞位置和信号肽预测结果表明,Lacc1含有信号肽,且为胞外分泌酶;Lacc1基因上游启动子区域包含1个TATAbox、3个金属响应元件(MRE)、5个氮因子结合位点(NIT2)、1个压力响应元件(STRE);在150μmol/L Cu2+的诱导下,Lacc1基因的表达在第6、8、10和14天分别上调了2.8、4.9、1.6和1.9倍,通过对Lacc1的启动子结构和诱导表达分析,推测灵芝漆酶注释基因Lacc1的诱导表达特性与启动子区域的金属响应元件等调控位点具有极大相关性。

To verify the function ofLaccl gene and investigate the relationship between the gene, promoter structure and transcription, expression for Ganoderma lucidum Laccase, we analyzed the amino acid structure of Laccl and upstream promoter region of putative laccase gene Laccl, whilst studied the putative laccase gene transcription in Ganoderma lucidum strain P9 induced by Cu2＋. Result of protein blast showed that Laccl had three Cu-oxidase conserved domains and multiple sequence alignments showed that Lacc 1 had the highest homology with lcc3-3 of polyporus ciliatus, which was 71%, and the highly conserved laccase signature sequence LI-L4 was also found in Laccl. The results of signal peptide prediction and sub-cellular location identification showed that Laccl is an extracellular （secreted） protein containing signal peptide. The prediction of upstream promoter region of gene Laccl showed 1 TATA box, 3 MRE （metal responsive element）, 5 NIT2 （nitrogen regulatory element） and 1 STRE （stress responsive element）. The expression of gene Laccl was up-regulated 2.8-fold, 4.9-fold, 1.6-fold and 1.9-fold at 6th day, 8th day, 10th day and 14th day after induction by 150 pmol/L Cu^2＋, respectively, in comparison to the reference condition. Via the analysis of Laccl sequenceand Cu〉 induced gene expression, the Cu^2＋ induced transcription ofLaccl gene was closely related to regulation motifs such as the responding elements for metals in the promoter region.