MOLECULAR ANALYSIS OF THE CORYNEBACTERIUM-GLUTAMICUM GDH GENE ENCODING GLUTAMATE-DEHYDROGENASE
MOLECULAR ANALYSIS OF THE CORYNEBACTERIUM-GLUTAMICUM GDH GENE ENCODING GLUTAMATE-DEHYDROGENASE
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DOI：
10.1111/j.1365-2958.1992.tb01474.x
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发表时间：
1992-02-01
影响因子：
3.6
通讯作者：
SAHM, H
中科院分区：
文献类型：
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作者：
BORMANN, ER;EIKMANNS, BJ;SAHM, H
通过与大肠杆菌 gdh 突变体 PA340 互补，分离出编码 NADP 依赖性谷氨酸脱氢酶 (GDH) 的谷氨酸棒杆菌 gdh 基因。 gdh 基因被亚克隆到大肠杆菌/C 中。谷氨酸穿梭载体pEKO并引入谷氨酸棒杆菌。相对于野生型 (1.8U mg Protein-1)，重组菌株表现出大约八倍高的 GDH 特异性活性 (15U mg Protein-1)。对野生型和重组谷氨酸棒杆菌菌株的生理学研究表明没有迹象表明 gdh 表达具有显着调节作用。确定了2082bp的DNA序列，包括gdh基因、5'-和3'-侧翼区域。该结构基因由1344bp组成，编码448个氨基酸残基的多肽(M(r)49152)，与报道的来自其他生物体的谷氨酸脱氢酶的氨基酸序列具有高达53.6%的同一性。 Northern印迹杂交显示出1.65kb的mRNA转录物，表明谷氨酸棒杆菌的gdh基因是单顺反子的。转录从位于 AUG 上游 284bp 的 G 残基开始，被认为是翻译起始密码子。
The Corynebacterium glutamicum gdh gene encoding NADP-dependent glutamate dehydrogenase (GDH) has been isolated by complementation of the Escherichia coli gdh mutant PA340. The gdh gene was subcloned into the E. coli/C. glutamicum shuttle vector pEKO and introduced into C. glutamicum. Recombinant strains showed approximately eightfold higher specific GDH activity (15U mg protein-1) relative to the wild type (1.8U mg protein-1). Physiological studies with wild-type and recombinant C. glutamicum strains revealed no indication of significant regulation of gdh expression. The DNA sequence of 2082bp, including the gdh gene, 5'-, and 3'-flanking regions, was determined. The structural gene consists of 1344bp and codes for a polypeptide of 448 amino acid residues (M(r) 49152) showing up to 53.6% identity with reported amino acid sequences of glutamate dehydrogenases from other organisms. Northern blot hybridization revealed a 1.65kb mRNA transcript, indicating that the gdh gene of C. glutamicum is monocistronic. Transcription occurred from a G residue located 284bp upstream of the AUG considered to be the translational initiation codon.