Mechanism for folate-independent aldolase reaction catalyzed by serine hydroxymethyltransferase
Mechanism for folate-independent aldolase reaction catalyzed by serine hydroxymethyltransferase
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DOI：
10.1111/j.1742-4658.2011.08443.x
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发表时间：
2012-02-01
期刊：
影响因子：
5.4
通讯作者：
Igarashi, Yasuo
中科院分区：
文献类型：
--
作者：
Chiba, Yoko;Terada, Tohru;Igarashi, Yasuo
丝氨酸羟甲基转移酶在缺乏四氢叶酸的情况下催化β -羟氨基酸裂解成甘氨酸和醛。该酶接受各种β -羟氨基酸作为该反应的底物。反应速率取决于C原子上的取代基和立体化学：红基取代基和-苯基取代基分别优于三基取代基和-甲基取代基。虽然提出了几种机制，但决定底物偏好的因素仍不清楚。我们首先进行了量子力学计算来评估反应机制的有效性。结果表明，从β -羟基上提取质子开始的反醛醇机制是合理的。这也表明CaC β键的裂解是速率限制步骤。接下来，我们测量了四种代表性底物的速率常数与温度的关系，并从Arrhenius图中计算了活化能和指前因子。红血球形式的活化能低于三种形式的活化能。苯基取代基降低了三价形式的活化能，而在红价形式中，它没有改变活化能，但增加了指前因子。我们提出了一个统一的模型，通过结合理论和实验结果来解释取代基和立体化学偏好的起源。还讨论了嗜热菌中不依赖四氢叶酸活性的可能生物学作用。
Serine hydroxymethyltransferase catalyzes the cleavage of beta-hydroxyamino acids into glycine and aldehydes in the absence of tetrahydrofolate. The enzyme accepts various beta-hydroxyamino acids as the substrate of this reaction. The reaction rate varies depending on the substituent and stereochemistry at the C beta atom: the erythro forms and the beta-phenyl substituent are preferred over the threo forms and the beta-methyl substituent, respectively. Although several mechanisms have been proposed, what determines the substrate preference remains unclear. We first performed quantum mechanical calculations to assess the validity of the reaction mechanisms. The results indicate that the retro-aldol mechanism starting with abstraction of the proton from the beta-hydroxyl group is plausible. This also suggests that CaC beta bond cleavage is the rate-limiting step. We next measured the dependence of the rate constants on temperature with four representative substrates and calculated the activation energies and pre-exponential factors from the Arrhenius plots. The activation energies of the erythro forms were lower than those of the threo forms. The beta-phenyl substituent lowered the activation energy in the threo form, whereas it did not alter the activation energy but increased the pre-exponential factor in the erythro form. We present a unified model to explain the origin of the substituent and stereochemical preferences by combining the theoretical and experimental results. A possible biological role of the tetrahydrofolate-independent activity in thermophiles is also discussed.