Duan Xiaojie, Liu Yu, You Xin, Jiang Zhengqiang, Yang Shaoxiang, Yang Shaoqing
Beijing Advanced Innovation Center for Food Nutrition and Human Health, China Agricultural University, Beijing, 100083 China.
College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, 100083 China.
Biotechnol Biofuels. 2017 Sep 19;10:223. doi: 10.1186/s13068-017-0912-z. eCollection 2017.
Butyl butyrate has been considered as a promising fuel source because it is a kind of natural ester which can be converted from renewable and sustainable lignocellulosic biomass. Compared with the conventional chemical methods for butyl butyrate production, the enzymatic approach has been demonstrated to be more attractive, mainly owing to the mild reaction conditions, high specificity, low energy consumption, and environmental friendliness. Cutinases play an important role in the butyl butyrate production process. However, the production level of cutinases is still relatively low. Thus, to identify novel cutinases suitable for butyl butyrate synthesis and enhance their yields is of great value in biofuel industry.
A novel cutinase gene () was cloned from a thermophilic fungus and expressed in . The highest cutinase activity of 12, 536 U/mL was achieved in 5-L fermentor, which is by far the highest production for a cutinase. McCut was optimally active at pH 8.0 and 45 °C. It exhibited excellent stability within the pH range of 3.0-10.5 and up to 75 °C. The cutinase displayed broad substrate specificity with the highest activity towards -nitrophenyl butyrate and tributyrin. It was capable of hydrolyzing cutin, polycaprolactone, and poly(butylene succinate). Moreover, McCut efficiently synthesized butyl butyrate with a maximum esterification efficiency of 96.9% at 4 h. The overall structure of McCut was resolved as a typical α/β-hydrolase fold. The structural differences between McCut and cutinase in groove and loop provide valuable information for redesign of McCut. These excellent features make it useful in biosynthesis and biodegradation fields.
A novel cutinase from was identified and characterized for the first time. High-level expression by is by far the highest for a cutinase. The enzyme exhibited excellent stability and high esterification efficiency for butyl butyrate production, which may make it a good candidate in biofuel and chemical industries.
丁酸丁酯被认为是一种很有前景的燃料来源，因为它是一种天然酯类，可由可再生且可持续的木质纤维素生物质转化而来。与传统的丁酸丁酯化学生产方法相比，酶法已被证明更具吸引力，主要是由于其反应条件温和、特异性高、能耗低且环境友好。角质酶在丁酸丁酯生产过程中发挥着重要作用。然而，角质酶的产量仍然相对较低。因此，鉴定适合丁酸丁酯合成的新型角质酶并提高其产量在生物燃料工业中具有重要价值。
从嗜热真菌中克隆出一个新型角质酶基因（）并在中表达。在5升发酵罐中实现了最高12536 U/mL的角质酶活性，这是迄今为止角质酶的最高产量。McCut在pH 8.0和45℃时活性最佳。它在pH 3.0 - 10.5范围内以及高达75℃时表现出优异的稳定性。该角质酶表现出广泛的底物特异性，对丁酸对硝基苯酯和三丁酸甘油酯活性最高。它能够水解角质、聚己内酯和聚丁二酸丁二醇酯。此外，McCut能高效合成丁酸丁酯，在4小时时最大酯化效率达96.9%。McCut的整体结构解析为典型的α/β水解酶折叠。McCut与角质酶在凹槽和环区域的结构差异为McCut的重新设计提供了有价值的信息。这些优异特性使其在生物合成和生物降解领域有用。
首次鉴定并表征了一种来自的新型角质酶。通过实现的高水平表达是迄今为止角质酶的最高水平。该酶在丁酸丁酯生产中表现出优异的稳定性和高酯化效率，这可能使其成为生物燃料和化学工业中的良好候选者。
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