Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center in Shreveport, 1501 Kings Highway, Shreveport, Louisiana 71130, USA.
Biochemistry. 2011 Aug 23;50(33):7132-45. doi: 10.1021/bi200989h. Epub 2011 Jul 26.
Charcot-Marie-Tooth disorder (CMT) is the most common inherited peripheral neuropathy, afflicting 1 in every 2500 Americans. One form of this disease, Dominant Intermediate Charcot-Marie-Tooth disorder type C (DI-CMTC), is due to mutation of the gene encoding the cytoplasmic tyrosyl-tRNA synthetase (TyrRS). Three different TyrRS variants have been found to give rise to DI-CMTC: replacing glycine at position 41 by arginine (G41R), replacing glutamic acid at position 196 by lysine (E196K), and deleting amino acids 153-156 (Δ(153-156)). To test the hypothesis that DI-CMTC is due to a defect in the ability of tyrosyl-tRNA synthetase to catalyze the aminoacylation of tRNA(Tyr), we have expressed each of these variants as recombinant proteins and used single turnover kinetics to characterize their abilities to catalyze the activation of tyrosine and its subsequent transfer to the 3' end of tRNA(Tyr). Two of the variants, G41R and Δ(153-156), display a substantial decrease in their ability to bind tyrosine (>100-fold). In contrast, the E196K substitution does not significantly affect the kinetics for formation of the tyrosyl-adenylate intermediate and actually increases the rate at which the tyrosyl moiety is transferred to tRNA(Tyr). The observation that the E196K substitution does not decrease the rate of catalysis indicates that DI-CMTC is not due to a catalytic defect in tyrosyl-tRNA synthetase.
腓骨肌萎缩症（CMT）是最常见的遗传性周围神经病，每 2500 名美国人中就有 1 人患有这种疾病。这种疾病的一种形式，即显性中间型腓骨肌萎缩症 C 型（DI-CMTC），是由于编码细胞质酪氨酸-tRNA 合成酶（TyrRS）的基因突变引起的。已经发现三种不同的 TyrRS 变体导致 DI-CMTC：甘氨酸第 41 位被精氨酸取代（G41R）、谷氨酸第 196 位被赖氨酸取代（E196K）和缺失氨基酸 153-156（Δ（153-156））。为了检验 DI-CMTC 是由于 TyrRS 催化 tRNA(Tyr)氨酰化的能力缺陷的假设，我们已经将这些变体中的每一个都表达为重组蛋白，并使用单次转换动力学来表征它们催化酪氨酸激活及其随后转移到 tRNA(Tyr)3'端的能力。其中两种变体，G41R 和 Δ（153-156），显示出其结合酪氨酸的能力显著下降（超过 100 倍）。相比之下，E196K 取代不会显著影响形成酪氨酸腺苷酸中间物的动力学，实际上增加了酪氨酸部分转移到 tRNA(Tyr)的速率。E196K 取代不会降低催化速率的观察表明，DI-CMTC 不是由于 TyrRS 催化缺陷引起的。
