中文摘要：

【目的】克隆猴头菌CB1锰过氧化物酶（Mn P）基因,用于分析He-mnp1基因及蛋白序列、基因的结构与功能,为研究猴头菌Mn Ps基因功能、转录调控和构建优良工程菌株提供参考。【方法】根据Gen Bank中已报道的白腐菌Mn Ps基因c DNA序列的保守区域设计引物,采用简并PCR、逆转录RT-PCR、c DNA末端的快速扩增（RACE）等方法扩增出全长c DNA基因序列,命名为He-mnp1（Gen Bank登录号为HM116841.3）,并对其猴头菌He-mnp1基因进行生物信息学的分析。通过NCBI数据库进行BLAST同源搜索;ORF Finder查找该基因的完整开放阅读框（ORF）;利用Expasy数据库和Bio Edit软件预测He-mnp1蛋白质的理化特性及氨基酸的组成;同时进行亲/疏水性及跨膜区的分析;采用Signal P 4.1软件进行蛋白信号肽的预测;并利用Clustal W和MEGA 5.1软件对He-mnp1蛋白序列同源性比对和构建白腐菌Mn Ps系统发育树;利用数据库Conserved Domain Database（CDD）蛋白保守结构域的预测,查看He-mnp1血红素、基质及锰、钙等结合位点;采用Predict Protein软件和SWISS-MODEL软件进行He-mnp1蛋白二级结构的预测和同源三维建模。【结果】该基因He-mnp1的c DNA全长1 279 bp,完整开放阅读框1 080 bp,起始密码子ATG,终止密码子TAA,5＇端非翻译区有68个核苷酸,3＇端非翻译区有131个核苷酸,编码蛋白359 aa;生物信息学分析表明,猴头菌He-mnp1氨基酸组成中丙氨酸含量最高,无酪氨酸,分子量为38.18 k Da,等电点为4.35,He-mnp1的蛋白有明显的亲水区,存在81-105、121-141间有2处疏水区域,此蛋白为亲水蛋白。He-mnp1蛋白质多肽前体包含1个18 aa的信号肽及1个5 aa的中间前导短肽。【结论】蛋白系统进化分析表明He-mnp1分布在第2组群,与平菇侧耳、冬生多孔菌、变色栓菌的Mn Ps亲缘关系最为接近。He-mnp1基因存在1个保守结构域,分析显示为ClassⅡ类的真菌血红素过氧化物酶蛋白家族。He-mnp1蛋白?

英文摘要：

[Objective]The cDNA gene sequences of Manganese peroxidase （ MnP） were isolated from H. erinaceum CB1,and used for analyzing structure and function of the He-mnp1.[Method]Degenerate primers were designed according to conservative domain of white-rot fungi MnPs gene cDNA sequences reported in GenBank,the full-length cDNA gene sequence was obtained by using the methods of PCR,Reverse transcription-PCR and Rapid Amplification of cDNA Ends （RACE） and named as He-mnp1（GenBank No. HM116841. 3）,and the bioinformatics of He-mnp1 gene were analyzed. BLAST homology search was conducted through the NCBI database; ORF Finder was used to look up the complete open reading frame of the gene; The Expasy database and BioEdit software were used to predict physicochemical properties and amino acid composition of He-mnp1 protein,and analyze the hydrophilicity/hydrophobicity and transmembrane region;The SignalP 4. 1 software was used to predict protein signal peptide; The clustal W with MEGA 5. 1 software was adopted to complete the He-mnp1 protein sequence homology alignment and to construct the phylogenetic trees of white rot fungi MnPs,respectively. The CDD database was used to predict protein conserved domains,and check the He-mnp1 heme,substrate and manganese,calcium binding site etc. The PredictProtein software and SWISS-MODEL software were used to complete the He-mnp1 protein secondary structure prediction and to construct homologous 3D modeling,respectively.[Result]The full-length cDNA of He-mnp1 was 1 279 bp,the ORF of 1 080 bp with starting codon of ATG and stopping codon of TAA,including 5＆#39;UTR of 68 bps and 3＆#39;UTR of 131 bps and encoded 359 amino acids. Bioinformatics analysis showed that the He-mnp1 protein has the highest content of Ala,without Tyr,and the Mw 38. 18 is kDa,with the pI of 4. 35 . The He-nmp1 protein has an obvious hydrophilic region and two hydrophobic regions in the area of 81 -105 and 121 -141,and belongs to hydrophilic protein. He-mnp1 protein precursor polypeptides consists of a