Evaluation of fluoropyruvate as nucleophile in reactions catalysed by N-acetyl neuraminic acid lyase variants: scope, limitations and stereoselectivity.
Evaluation of fluoropyruvate as nucleophile in reactions catalysed by N-acetyl neuraminic acid lyase variants: scope, limitations and stereoselectivity.
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DOI：
10.1039/c5ob02037a
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发表时间：
2016-01-07
影响因子：
3.2
通讯作者：
Nelson A
中科院分区：
文献类型：
--
作者：
Stockwell J;Daniels AD;Windle CL;Harman TA;Woodhall T;Lebl T;Trinh CH;Mulholland K;Pearson AR;Berry A;Nelson A
报道了氟丙酮酸与醛在醛缩酶催化下反应的立体化学过程。研究了N-乙酰神经氨酸裂解酶（NAL）变体对氟丙酮酸作为供体的反应的催化作用。在动力学控制下，野生型酶催化氟丙酮酸和N-乙酰甘露糖胺之间的反应，得到90：10比例的（3R，4 R）-和（3S，4 R）-构型产物;在延长的反应时间后，发生平衡，得到这些产物的30：70混合物。还研究了E192 N、E192 N/T167 V/S208 V和E192 N/T167 G NAL变体催化的一系列底物的反应的效率和立体选择性。使用氟丙酮酸盐和（2 R，3S）-或（2S，3R）-2，3-二羟基-4-氧代-N，N-二丙基丁酰胺作为底物，可以获得四种可能的非对映体产物中的三种;对于每种产物，测定异头和吡喃糖/呋喃糖形式的比率。对S.测定了与氟丙酮酸复合的金黄色葡萄球菌NAL，有助于合理化C-C键形成的立体化学结果。
The stereochemical course of aldolase-catalysed reaction between fluoropyruvate and aldehydes is described. The catalysis of reactions involving fluoropyruvate as donor by N-acetyl neuraminic acid lyase (NAL) variants was investigated. Under kinetic control, the wild-type enzyme catalysed the reaction between fluoropyruvate and N-acetyl mannosamine to give a 90 : 10 ratio of the (3R,4R)- and (3S,4R)-configured products; after extended reaction times, equilibration occurred to give a 30 : 70 mixture of these products. The efficiency and stereoselectivity of reactions of a range of substrates catalysed by the E192N, E192N/T167V/S208V and E192N/T167G NAL variants were also studied. Using fluoropyruvate and (2R,3S)- or (2S,3R)-2,3-dihydroxy-4-oxo-N,N-dipropylbutanamide as substrates, it was possible to obtain three of the four possible diastereomeric products; for each product, the ratio of anomeric and pyranose/furanose forms was determined. The crystal structure of S. aureus NAL in complex with fluoropyruvate was determined, assisting rationalisation of the stereochemical outcome of C–C bond formation.