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首页> 外文期刊>Journal of bacteriology >Alpha-amylase genes (amyR2 and amyE+) from an alpha-amylase-hyperproducing Bacillus subtilis strain: molecular cloning and nucleotide sequences.
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Alpha-amylase genes (amyR2 and amyE+) from an alpha-amylase-hyperproducing Bacillus subtilis strain: molecular cloning and nucleotide sequences.

机译：来自高产α-淀粉酶的枯草芽孢杆菌菌株的α-淀粉酶基因（amyR2和amyE +）：分子克隆和核苷酸序列。

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摘要

amyR2, amyE+, and aroI+ alleles from an alpha-amylase-hyperproducing strain, Bacillus subtilis NA64, were cloned in temperate B. subtilis phage p11, and the amyR2 and amyE+ genes were then recloned in plasmid pUB110, which was designated pTUB4. The order of the restriction sites, ClaI-EcoRI-PstI-SalI-SmaI, found in the DNA fragment carrying amyR2 and amyE+ from the phage genome was also found in the 2.3-kilobase insert of pTUB4. Approximately 2,600 base pairs of the DNA nucleotide sequence of the amyR2 and amyE+ gene region in pTUB4 were determined. Starting from an ATG initiator codon, an open reading frame was composed of a total 1,776 base pairs (592 amino acids). Among the 1,776 base pairs, 1,674 (558 amino acids) were found in the cloned DNA fragment, and 102 base pairs (34 amino acids) were in the vector pUB110 DNA. The COOH terminal region of the alpha-amylase of pTUB4 was encoded in pUB110. The electrophoretic mobility in a 7.5% polyacrylamide gel of the alpha-amylase was slightly faster than that of the parental alpha-amylases. The NH2 termination portion of the gene encoded a 41-amino acid-long signal sequence (Ohmura et al., Biochem. Biophys. Res. Commun. 112:687-683, 1983). The DNA sequence of the mature extracellular alpha-amylase, a potential RNA polymerase recognition site and Pribnow box (TTGATAGAGTGATTGTGATAATTTAAAAT), and an AT-rich inverted repeat structure which has free energy of -8.2 kcal/mol (-34.3 kJ/mol) were identified. The AT-rich inverted repeat structure seemed to correspond to the hyperproducing character. The nucleotide sequence around the region was quite different from the promoter region of the B. subtilis 168 alpha-amylase gene which was cloned in the Escherichia coli vector systems.
机译：将来自α-淀粉酶高产菌株枯草芽孢杆菌NA64的amyR2，amyE +和aroI +等位基因克隆到温和的枯草芽孢杆菌噬菌体p11中，然后将amyR2和amyE +基因重新克隆到质粒pUB110中，命名为pTUB4。在来自噬菌体基因组的携带amyR2和amyE +的DNA片段中发现的限制性酶切位点ClaI-EcoRI-PstI-SalI-SmaI的顺序也在pTUB4的2.3碱基碱基插入物中发现。确定了pTUB4中amyR2和amyE +基因区域的DNA核苷酸序列的大约2,600个碱基对。从ATG起始密码子开始，开放阅读框由总共1,776个碱基对（592个氨基酸）组成。在1,776个碱基对中，在克隆的DNA片段中发现了1,674个（558个氨基酸），在载体pUB110 DNA中发现了102个碱基对（34个氨基酸）。在pUB110中编码了pTUB4的α-淀粉酶的COOH末端区域。在7.5％聚丙烯酰胺凝胶中，α-淀粉酶的电泳迁移率略快于亲本α-淀粉酶。该基因的NH 2终止部分编码一个41个氨基酸长的信号序列（Ohmura等，Biochem.Biophys.Res.Commun.112：687-683，1983）。成熟细胞外α-淀粉酶的DNA序列，潜在的RNA聚合酶识别位点和Pribnow盒（TTGATAGAGTGATTGTGATAATTTAAAAT）以及自由能为-8.2 kcal / mol（-34.3 kJ / mol）的富含AT的反向重复结构是确定。富含AT的反向重复结构似乎对应于高产特征。该区域周围的核苷酸序列与枯草芽孢杆菌168α-淀粉酶基因的启动子区域完全不同，枯草芽孢杆菌168α-淀粉酶基因被克隆到大肠杆菌载体系统中。

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