Tsai Shau-Wei, Chen Chun-Chi, Yang Hung-Shien, Ng I-Son, Chen Teh-Liang
Institute of Biochemical and Biomedical Engineering, Chang Gung University, Kwei-Shan Tao-Yuan, 33302, Taiwan.
Biochim Biophys Acta. 2006 Aug;1764(8):1424-8. doi: 10.1016/j.bbapap.2006.07.001. Epub 2006 Jul 14.
In comparison with the biocatalyst engineering and medium engineering approaches, very few examples have been reported on using the substrate engineering approach such as substrate-assisted catalysis (SAC) for naturally occurring or engineered lipases and serine proteases to improve the enzyme activity and enantioselectivity. By employing lipase-catalyzed hydrolysis of (R,S)-naproxen esters in water-saturated isooctane as the model system, we demonstrate the proton shuttle device to the leaving alcohol of the substrate as a new means of SAC to effectively improve the lipase activity or enantioselectivity. The result cannot only provide a strong evidence for the rate-limiting proton transfer for the bond-breaking of tetrahedron intermediate of the acylation step, but also sheds light for performing the hydrolysis, transesterification or aminolysis in organic solvents for the ester substrate that originally lipases cannot catalyze, but now can after introducing the device.
与生物催化剂工程和介质工程方法相比，利用底物工程方法（如底物辅助催化，SAC）来提高天然存在或工程化脂肪酶和丝氨酸蛋白酶的酶活性和对映选择性的报道实例非常少。通过以水饱和异辛烷中脂肪酶催化（R,S）-萘普生酯的水解作为模型体系，我们证明了向底物离去醇的质子穿梭装置是一种有效的SAC新手段，可有效提高脂肪酶活性或对映选择性。该结果不仅为酰化步骤四面体中间体断键的限速质子转移提供了有力证据，也为在有机溶剂中对原本脂肪酶无法催化但引入该装置后现在可以催化的酯底物进行水解、酯交换或氨解提供了思路。
