【作者】 王更先； 牛艳山； 司马杨虎； 徐世清；

【Author】 WANG Geng-Xian1,ZHANG Sheng-Xiang1,2,LIU Yan-Shan1,SIMA Yang-Hu1and XU Shi-Qing1(1.National Engineering Laboratory of Modern Silk,Department of Applied Biology,Medical College,Soochow University,Suzhou,Jiangsu 215123,China;2.College of Forestry,Shandong Agricultural University,Taian,Shandong 271018,China)

【机构】 现代丝绸国家工程实验室苏州大学医学部应用生物学系； 山东农业大学林学院；

【摘要】 脱氧羟腐胺赖氨酸合酶和脱氧羟腐胺赖氨酸羟化酶是催化真核翻译起始因子序列中羟腐胺赖氨酸合成的两个酶。羟腐胺赖氨酸的合成是真核翻译起始因子活化成熟的标志,对真核细胞的存活及增殖具有重要作用。本文利用生物信息学、RACE和RT-PCR等方法克隆了家蚕脱氧羟腐胺赖氨酸合酶和脱氧羟腐胺赖氨酸羟化酶的同源基因BmDHS和BmDOHH,分析了基因结构和表达谱。BmDHS和BmDOHH的cDNA全长分别为1311bp和1874bp。BmDHS只有一个外显子,没有内含子,包含最大ORF框为1116bp,编码371个氨基酸,分子量为41.11KD,等电点为5.84。BmDOHH由4个外显子和3个内含子构成,包含最大ORF框为915bp,编码304个氨基酸,分子量为34.30kD,等电点为4.86。BmDHS编码的蛋白质在47-361氨基酸残基位置为脱氧羟腐胺赖氨酸合酶的保守区域;BmDOHH编码的蛋白质分别在23-52,54-83,87-116,177-206,208-237,241-270氨基酸残基位置存在E-ZtypeHEAT重复区域。BmDHS和BmDOHH所编码的蛋白质与人、果蝇等生物均具有55%以上的蛋白质同源性,保守区域高度一致。系统发育树分析结果显示DHS和DOHH在进化上存在物种特异性。表达谱分析表明BmDHS和BmDOHH在所调查的组织中均没有组织和时空特异性。更多还原

【Abstract】 Deoxyhypusine snyhtase(DHS) and deoxyhypusine hydroxylase(DOHH) are the two enzymes that catalyze the synthesis of hypusine within eukaryotic initiation factor 5A(eIF5A).Synthesis of hypusine is essential for the function of eIF5A in eukaryotic cell proliferation and survival.Here we described the cloning and expression of two full length cDNAs,encoding respectively DHS-like protein and DOHH-like protein from Bombyx mori by using the methods of bioinformatics,RACE and RT-PCR technology,named as BmDHS and BmDOHH.Sequencing results indicate that they are 1311 bp and 1874 bp in length including complete open reading frame(ORF) 1116 bp and 915 bp,which encode 371 amino acids(molecular weight is about 41.11 kD and isoelectric point is 5.84) and 304 amino acids(molecular weight is about 34.30 kD and isoelectric point is 4.86),respectively.BmDHS contains only one exon,and BmDOHH contains 4 exons and 3 introns.The deduced amino acid sequence of BmDHS contains a deoxyhypusine synthase domain from 47 to 361 amino acid residues,and the deduced amino acid sequence of BmDOHH contains six E-Z type HEAT repeat domains(23-52,54-83,87-116,177-206,208-237,241-270).Compared to DHS and DOHH amino acid sequences from other species,such as Homo sapiens,Drosophila melanogaster,both silkworm DHS protein and DOHH protein have more than 55% identity.The conservative regions are very similar with each other.The phylogenetic tree analysis indicated that not only DHS but also DOHH from different species has genus-specific features.The expressions of BmDHS and BmDOHH are no tissue and stage specific in our tested samples.更多还原