NADP(+)-Specific Isocitrate Dehydrogenase from Oleaginous Yeast Yarrowia lipolytica CLIB122: Biochemical Characterization and Coenzyme Sites Evaluation
文献类型:期刊论文
作者 | Li X1; Wang P1; Ge YD1; Wang W2; Abbas A3; Zhu GP[*]1 |
刊名 | APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
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出版日期 | 2013 |
卷号 | 17期号:2页码:403-416 |
关键词 | Isocitrate dehydrogenase Yarrowia lipolytica Kinetics Coenzyme specificity Site-directed mutagenesis |
通讯作者 | gpz1996@yahoo.com |
合作状况 | 其它 |
英文摘要 | NADP(+)-dependent isocitrate dehydrogenase from Yarrowia lipolytica CLIB122 (YlIDP) was overexpressed and purified. The molecular mass of YlIDP was estimated to be about 81.3 kDa, suggesting its homodimeric structure in solution. YlIDP was divalent cation dependent and Mg2+ was found to be the most favorable cofactor. The purified recombinant YlIDP displayed maximal activity at 55 A degrees C and its optimal pH for catalysis was found to be around 8.5. Heat inactivation studies revealed that the recombinant YlIDP was stable below 45 A degrees C, but its activity dropped quickly above this temperature. YlIDP was absolutely dependent on NADP(+) and no NAD-dependent activity could be detected. The K (m) values displayed for NADP(+) and isocitrate were 59 and 31 mu M (Mg2+), 120 mu M and 58 mu M (Mn2+), respectively. Mutant enzymes were constructed to tentatively alter the coenzyme specificity of YlIDP. The K (m) values for NADP(+) of R322D mutant was 2,410 mu M, being about 41-fold higher than that of wild type enzyme. NAD(+)-dependent activity was detected for R322D mutant and the K (m) and k (cat) values for NAD(+) were 47,000 mu M and 0.38 s(-1), respectively. Although the R322D mutant showed low activity with NAD(+), it revealed the feasibility of engineering an eukaryotic IDP to a NAD(+)-dependent one. |
收录类别 | SCI |
资助信息 | This research was supported by funds from the National High Technology Research and Development Program (“863” Program: 2012AA02A708), the National Natural Science Foundation of China (31170005 and 30870062), Specialized Research Fund for the Doctoral Program of Higher Education of China (20113424110004), the Fund of State Key Laboratory of Genetics Resources and Evolution from Kunming Institute of Zoology (Chinese Academy of Sciences, CAS) (GREKF11-07), and Anhui Provincial Natural Science Foundation (1208085QC52 and 1308085QC67). |
语种 | 英语 |
WOS记录号 | WOS:000324110500011 |
公开日期 | 2013-10-11 |
源URL | [http://159.226.149.42:8088/handle/152453/7693] ![]() |
专题 | 昆明动物研究所_基因起源组 昆明动物研究所_遗传资源与进化国家重点实验室 |
作者单位 | 1.Institute of Molecular Biology and Biotechnology and Anhui Provincial Key Laboratory of the Conservation and Exploitation of Biological Resources, Anhui Normal University, Wuhu 241000, Anhui, China 2.State Key Laboratory of Genetics Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650223, Yunnan, China 3.College of Life Science and Technology, Xinjiang University, Urumqi 830046, Xinjiang, China |
推荐引用方式 GB/T 7714 | Li X,Wang P,Ge YD,et al. NADP(+)-Specific Isocitrate Dehydrogenase from Oleaginous Yeast Yarrowia lipolytica CLIB122: Biochemical Characterization and Coenzyme Sites Evaluation[J]. APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY,2013,17(2):403-416. |
APA | Li X,Wang P,Ge YD,Wang W,Abbas A,&Zhu GP[*].(2013).NADP(+)-Specific Isocitrate Dehydrogenase from Oleaginous Yeast Yarrowia lipolytica CLIB122: Biochemical Characterization and Coenzyme Sites Evaluation.APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY,17(2),403-416. |
MLA | Li X,et al."NADP(+)-Specific Isocitrate Dehydrogenase from Oleaginous Yeast Yarrowia lipolytica CLIB122: Biochemical Characterization and Coenzyme Sites Evaluation".APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY 17.2(2013):403-416. |
入库方式: OAI收割
来源:昆明动物研究所
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