Second-Shell Amino Acid R266 Helps Determine N-Succinylamino Acid Racemase Reaction Specificity in Promiscuous N-Succinylamino Acid Racemase/o-Succinylbenzoate Synthase Enzymes.
Second-Shell Amino Acid R266 Helps Determine N-Succinylamino Acid Racemase Reaction Specificity in Promiscuous N-Succinylamino Acid Racemase/o-Succinylbenzoate Synthase Enzymes.
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DOI：
10.1021/acs.biochem.1c00627
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发表时间：
2021-12-21
期刊：
影响因子：
2.9
通讯作者：
Glasner ME
中科院分区：
文献类型：
--
作者：
Truong DP;Rousseau S;Machala BW;Huddleston JP;Zhu M;Hull KG;Romo D;Raushel FM;Sacchettini JC;Glasner ME
催化混杂性是在与天然生物反应相同的活性位点催化非生物反应的巧合能力。一些证据表明，催化混杂在新的酶功能的进化中发挥作用。因此，研究催化混杂性可以帮助确定使酶易于进化新功能的结构特征。本研究确定了一个潜在的预适应残基的混杂N-琥珀酰氨基酸消旋酶/邻琥珀酰苯甲酸合酶（NSAR/OSBS）酶拟无枝酸菌属物种T-1-60。该酶属于OSBS家族的分支，该家族包括许多催化混杂的NSAR/OSBS酶。R266在NSAR/OSBS亚家族的所有成员中是保守的。然而，在其他OSBS亚家族中，同源位置通常是疏水的，其酶缺乏NSAR活性。第二壳层氨基酸R266与催化剂酸碱K263接近，但不与底物接触，表明R266可能影响催化机理。将拟无枝酸菌NSAR/OSBS中的R266突变为谷氨酰胺显著降低了NSAR活性，但适度降低了OSBS活性。这是由于底物和一般酸/碱催化剂K263之间的质子交换速率降低了1000倍。这种突变对OSBS反应的危害较小，因为K263与OSBS底物和/或中间体形成阳离子-π相互作用，而不是充当一般的酸/碱催化剂。总之，这些数据解释了R266如何有助于NSAR反应特异性，并且可能是NSAR活性进化的必要预适应。
Catalytic promiscuity is the coincidental ability to catalyze non-biological reactions in the same active site as the native biological reaction. Several lines of evidence show that catalytic promiscuity plays a role in the evolution of new enzyme functions. Thus, studying catalytic promiscuity can help identify structural features that predispose an enzyme to evolve new functions. This study identifies a potentially pre-adaptive residue in a promiscuous N-succinylamino acid racemase/o-succinylbenzoate synthase (NSAR/OSBS) enzyme from Amycolatopsis sp. T-1–60. This enzyme belongs to a branch of the OSBS family which includes many catalytically promiscuous NSAR/OSBS enzymes. R266 is conserved in all members of the NSAR/OSBS subfamily. However, the homologous position is usually hydrophobic in other OSBS subfamilies, whose enzymes lack NSAR activity. The second-shell amino acid R266 is close to the catalytic acid/base K263, but it does not contact the substrate, suggesting that R266 could affect the catalytic mechanism. Mutating R266 to glutamine in Amycolatopsis NSAR/OSBS profoundly reduces NSAR activity, but moderately reduces OSBS activity. This is due to a 1000-fold decrease in the rate of proton exchange between the substrate and the general acid/base catalyst K263. This mutation is less deleterious for the OSBS reaction because K263 forms a cation-π interaction with the OSBS substrate and/or the intermediate, rather than acting as a general acid/base catalyst. Together, the data explain how R266 contributes to NSAR reaction specificity and was likely an essential preadaptation for the evolution of NSAR activity.
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影响因子：
9.8
作者：
Jack BR;Meyer AG;Echave J;Wilke CO
通讯作者：
Wilke CO
通过同源性建模和虚拟筛选引导的二肽分配酶酶促功能的发现。
DOI：
10.1016/j.str.2008.08.015
发表时间：
2008-11-12
期刊：
Structure (London, England : 1993)
影响因子：
--
作者：
Kalyanaraman C;Imker HJ;Fedorov AA;Fedorov EV;Glasner ME;Babbitt PC;Almo SC;Gerlt JA;Jacobson MP
通讯作者：
Jacobson MP
影响因子：
2.9
作者：
Blaisse, Michael R.;Fu, Beverly;Chang, Michelle C. Y.
通讯作者：
Chang, Michelle C. Y.
影响因子：
4.6
作者：
Golden E;Yu LJ;Meilleur F;Blakeley MP;Duff AP;Karton A;Vrielink A
通讯作者：
Vrielink A
影响因子：
2.9
作者：
Brodkin HR;Novak WR;Milne AC;D'Aquino JA;Karabacak NM;Goldberg IG;Agar JN;Payne MS;Petsko GA;Ondrechen MJ;Ringe D
通讯作者：
Ringe D