Single arginine mutation in two yeast isocitrate dehydrogenases: biochemical characterization and functional implication.
Single arginine mutation in two yeast isocitrate dehydrogenases: biochemical characterization and functional implication.
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两种酵母异柠檬酸脱氢酶中的单一精氨酸突变：生化特征和功能含义。
DOI：
10.1371/journal.pone.0115025
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Zhu G
中科院分区：
文献类型：
--
作者：
Song P;Wei H;Cao Z;Wang P;Zhu G
异柠檬酸脱氢酶（IDH）作为一种管家基因，引起了癌症专家的关注。人 IDH1 (HcIDH) 催化 Arg132 残基的突变消除了该酶的野生型异柠檬酸氧化活性，但赋予突变体将 α-酮戊二酸 (α-KG) 还原为 2-羟基戊二酸 (2-HG) 的能力。为了检查其他真核生物 IDH 中的类似突变是否会引起类似的影响，研究了两种酵母线粒体 IDH：酿酒酵母 NADP+-IDH1 (ScIDH1) 和解脂耶氏酵母 NADP+-IDH (YlIDH)。类似的 Arg 残基（ScIDH1 的 Arg148 和 YlIDH 的 Arg141）突变为 His。 ScIDH1 R148H和YlIDH R141H对于异柠檬酸的K m 值经测定分别比相应野生型酶的K m 值高2.4倍和2.2倍。 ScIDH1 R148H 和 YlIDH R141H 对异柠檬酸氧化的催化效率（k cat/K m）分别比野生型酶大幅降低了 227 倍和 460 倍。正如预期的那样，ScIDH1 R148H 和 YlIDH R141H 均获得了催化 α-KG 生成 2-HG 的新变体活性，并且使用气相色谱/飞行时间质谱 (GC/TOF-MS) 证实了 2-HG 的生成。动力学分析表明，ScIDH1 R148H 和 YlIDH R141H 对 α-KG 的亲和力分别比 HcIDH R132H 突变体高 5.2 倍和 3.3 倍。 ScIDH1 R148H和YlIDH R141H对α-KG的催化效率分别是HcIDH R132H突变体的5.5倍和4.5倍。由于HcIDH Arg132突变与肿瘤发生相关，本研究为进一步研究酵母体内IDH突变的生理作用提供了基础信息。
Isocitrate dehydrogenase (IDH), a housekeeping gene, has drawn the attention of cancer experts. Mutation of the catalytic Arg132 residue of human IDH1 (HcIDH) eliminates the enzyme's wild-type isocitrate oxidation activity, but confer the mutant an ability of reducing α-ketoglutarate (α-KG) to 2-hydroxyglutarate (2-HG). To examine whether an analogous mutation in IDHs of other eukaryotes could cause similar effects, two yeast mitochondrial IDHs, Saccharomyces cerevisiae NADP+-IDH1 (ScIDH1) and Yarrowia lipolytica NADP+-IDH (YlIDH), were studied. The analogous Arg residues (Arg148 of ScIDH1 and Arg141 of YlIDH) were mutated to His. The K m values of ScIDH1 R148H and YlIDH R141H for isocitrate were determined to be 2.4-fold and 2.2-fold higher, respectively, than those of the corresponding wild-type enzymes. The catalytic efficiencies (k cat/K m) of ScIDH1 R148H and YlIDH R141H for isocitrate oxidation were drastically reduced by 227-fold and 460-fold, respectively, of those of the wild-type enzymes. As expected, both ScIDH1 R148H and YlIDH R141H acquired the neomorphic activity of catalyzing α-KG to 2-HG, and the generation of 2-HG was confirmed using gas chromatography/time of flight-mass spectrometry (GC/TOF-MS). Kinetic analysis showed that ScIDH1 R148H and YlIDH R141H displayed 5.2-fold and 3.3-fold higher affinities, respectively, for α-KG than the HcIDH R132H mutant. The catalytic efficiencies of ScIDH1 R148H and YlIDH R141H for α-KG were 5.5-fold and 4.5-fold, respectively, of that of the HcIDH R132H mutant. Since the HcIDH Arg132 mutation is associated with the tumorigenesis, this study provides fundamental information for further research on the physiological role of this IDH mutation in vivo using yeast.
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DOI：
10.1126/science.1170944
发表时间：
2009-04-10
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Zhao S;Lin Y;Xu W;Jiang W;Zha Z;Wang P;Yu W;Li Z;Gong L;Peng Y;Ding J;Lei Q;Guan KL;Xiong Y
通讯作者：
Xiong Y
影响因子：
10.5
作者：
Chen C;Liu Y;Lu C;Cross JR;Morris JP 4th;Shroff AS;Ward PS;Bradner JE;Thompson C;Lowe SW
通讯作者：
Lowe SW
影响因子：
64.8
作者：
通讯作者：
--
影响因子：
64.8
作者：
Lu, Chao;Ward, Patrick S.;Kapoor, Gurpreet S.;Rohle, Dan;Turcan, Sevin;Abdel-Wahab, Omar;Edwards, Christopher R.;Khanin, Raya;Figueroa, Maria E.;Melnick, Ari;Wellen, Kathryn E.;O'Rourke, Donald M.;Berger, Shelley L.;Chan, Timothy A.;Levine, Ross L.;Mellinghoff, Ingo K.;Thompson, Craig B.
通讯作者：
Thompson, Craig B.
影响因子：
50.3
作者：
Ward PS;Patel J;Wise DR;Abdel-Wahab O;Bennett BD;Coller HA;Cross JR;Fantin VR;Hedvat CV;Perl AE;Rabinowitz JD;Carroll M;Su SM;Sharp KA;Levine RL;Thompson CB
通讯作者：
Thompson CB