Alternation in Substrate Specificity of Amino Acid Dehydrogenases based on the Structural Analyses
基于结构分析的氨基酸脱氢酶底物特异性的改变
基本信息
- 批准号:09460052
- 负责人:
- 金额:$ 8.45万
- 依托单位:
- 依托单位国家:日本
- 项目类别:Grant-in-Aid for Scientific Research (B)
- 财政年份:1997
- 资助国家:日本
- 起止时间:1997 至 1999
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
According to the structural analysis of glutamate dehydrogenase (GluDH) of Clostridium symbiosum, we studied the alternation of substrate specificity of phenylalanine dehydrogenase (PheDH) of Bacillus sphaericus to leucine dehydrogenase (LeuDH). We focused on residues which are not common in the amino acid dehydrogenases around the pocket for the substrate. 163A and 377V which are found in GluDH and LeuDH were changed to G and L in PheDH, respectively. The modified GluDH showed lower activity toward L-Phe, where as the activity toward aliphatic amino acids were increased.Opine dehydrogenase (ODH) from a soil isolate Arthrobacter sp. strain 1C showed similarity toward 40-kDa Protein, D-lysopine dehydrogenase, D-nopaline dehydrogenase found in plant crown gall. ODH was highly purified from E. coli transformant. Crystals of ODH, obtained in the presence or absence of co-factor and substrate, have been shown to diffract to beyond 1.8Å resolution. We carried out site-directed mutagenesis on these residues and other ones in both domains, and the results indicated that substitution of either of these six residues or Arg-143, Lys-156, His-222, or Tyr-280 impaired catalysis significantly. Steady-state kinetic interaction and catalysis. An opine analog N-[1-(S)-(carboxyl)ethyl]-(S)-phenylalanine competitively inhibited the wild type and most mutants with the exception of Arg143 mutants, Further, we observed a sulfate ion bound in the vicinity of His-202, Tyr-222, Arg-292 and Tyr-293 in the putative active site. We modeled a pyruvate in the sulfate binding site and a catalytic mechanism of ODH to resemble that of the 2-hydroxy acid dehydrogenases is proposed.
根据共生梭菌谷氨酸脱氢酶(GluDH)的结构分析,研究了球形芽孢杆菌苯丙氨酸脱氢酶(PheDH)对亮氨酸脱氢酶(LeuDH)底物专一性的改变。我们专注于在底物口袋周围的氨基酸脱氢酶中不常见的残基。在GluDH和LeuDH中发现的163 A和377 V在PheDH中分别变为G和L。土壤节杆菌1C菌株的冠瘿碱脱氢酶(ODH)与植物冠瘿中的40-kDa蛋白、D-赖氨酸脱氢酶、D-胭脂碱脱氢酶具有相似性。ODH是从E.大肠杆菌在存在或不存在辅因子和底物的情况下获得的ODH晶体已显示出超过1.8 μ m的分辨率。我们对这两个结构域中的这些残基和其他残基进行了定点突变,结果表明,这六个残基或Arg-143,Lys-156,His-222或Tyr-280中的任何一个的取代都会显着损害催化作用。稳态动力学相互作用与催化。冠瘿碱类似物N-[1-(S)-(羧基)乙基]-(S)-苯丙氨酸竞争性抑制野生型和除Arg 143突变体外的大多数突变体。此外,我们观察到在推定的活性位点的His-202、Tyr-222、Arg-292和Tyr-293附近结合有硫酸根离子。我们模拟了丙酮酸在硫酸盐结合位点和ODH的催化机制类似的2-羟基酸脱氢酶提出。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
K.L.Britton: "Crystallisation of Arthrobacter sp.Strain 1C opine dehydrogenase and its complex with NAD^+." Acta Cryst.D.54,. 124-126 (1998)
K.L.Britton:“节杆菌属菌株 1C 鸦片脱氢酶及其与 NAD^ 的复合物的结晶。”
- DOI:
- 发表时间:
- 期刊:
- 影响因子:0
- 作者:
- 通讯作者:
Y. Asano: "Screeing for novel amino acid dehdrogenases-soil of Toyama Prefecture as a source of microorganisms-(in Japanese)"Bulletin of toyama Prefctural University. 6. 101-106 (1996)
Y.浅野:“新型氨基酸脱氢酶的筛选-作为微生物来源的富山县土壤-(日语)”富山县立大学通报。
- DOI:
- 发表时间:
- 期刊:
- 影响因子:0
- 作者:
- 通讯作者:
Y. Gato and Y. Asano: "Opine dehydrogenase and secondary-amine dicarboxylic acids, in M. C. Flickinger and S. W. Drew (ed.), Encyclopedia of Bioprocess Technology"Fermentation, Biocatalysis, and Bioseparation, John Wiley & Sons, Inc., New York. 1851-1858
Y. Gato 和 Y. Asano:“Opine 脱氢酶和仲胺二羧酸,M. C. Flickinger 和 S. W. Drew(编辑),生物工艺技术百科全书”发酵、生物催化和生物分离,John Wiley
- DOI:
- 发表时间:
- 期刊:
- 影响因子:0
- 作者:
- 通讯作者:
Yasuhisa Asano: "Development of new microbial enzymes and their Application"Japanese Journal of Synthetic Organic chemistry. 57 (in Japanese). 1064-1072 (1999)
浅野康久:“新型微生物酶的开发及其应用”日本合成有机化学杂志。
- DOI:
- 发表时间:
- 期刊:
- 影响因子:0
- 作者:
- 通讯作者:
Y. Asano, and M. Tanetani: "Thermostable phenylalanine dehydrogenase from a mesophilic Microbacterium sp. strain DM-86-1."Arch Microbiol.. 169. 220-224 (1998)
Y. Asano 和 M. Tanetani:“来自嗜温微杆菌属菌株 DM-86-1 的耐热苯丙氨酸脱氢酶。”Arch Microbiol.. 169. 220-224 (1998)
- DOI:
- 发表时间:
- 期刊:
- 影响因子:0
- 作者:
- 通讯作者:
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
数据更新时间:{{ journalArticles.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ monograph.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ sciAawards.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ conferencePapers.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ patent.updateTime }}
ASANO Yasuhisa其他文献
ASANO Yasuhisa的其他文献
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
{{ truncateString('ASANO Yasuhisa', 18)}}的其他基金
Enzymatic synthesis of useful chemicals from nitrogen-containg substrates
从含氮底物酶法合成有用化学品
- 批准号:
23248015 - 财政年份:2011
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for Scientific Research (A)
Use of the enzymes in the microbial and plant "aldoxime-nitrile pathway"for chiral synthesis
利用微生物和植物“醛肟-腈途径”中的酶进行手性合成
- 批准号:
20380053 - 财政年份:2009
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Discovery of amino acid amide racemase and its use in the dynamic kinetic resolution of amino acid amides
氨基酸酰胺消旋酶的发现及其在氨基酸酰胺动态拆分中的应用
- 批准号:
18380061 - 财政年份:2006
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Comparative Biochemistry of the Aldoxime-Nitrile Pathway of Plant and Microbial Origins
植物和微生物来源的醛肟-腈途径的比较生物化学
- 批准号:
16380064 - 财政年份:2004
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Discovery of Microbial "Aldoxime-Nitrile Pathway" and Its Application to Nitrile Synthesis
微生物“醛肟-腈途径”的发现及其在腈合成中的应用
- 批准号:
13460045 - 财政年份:2001
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Reaction Mechanisms and Application of New Bacterial Enzymes
新型细菌酶的反应机理及应用
- 批准号:
08044218 - 财政年份:1996
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for international Scientific Research
Development of Novel Lyases and Their Application to the Synthesis of Optically Active Compounds
新型裂解酶的开发及其在光学活性化合物合成中的应用
- 批准号:
07660117 - 财政年份:1995
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Studies on the Novel Enzyme Phenylalanine Dehydrogenase.
新型酶苯丙氨酸脱氢酶的研究。
- 批准号:
05044138 - 财政年份:1993
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for international Scientific Research
相似海外基金
Is-beta-alanine-specific opine dehydrogenase present?
是否存在β-丙氨酸特异性阿片脱氢酶?
- 批准号:
08660258 - 财政年份:1996
- 资助金额:
$ 8.45万 - 项目类别:
Grant-in-Aid for Scientific Research (C)