A new lipase from Serratia marcescens SRICI-01 (Trx-SmL) was successfully overexpressed in Escherichia coli with thioredoxin (Trx) fusion tag. Intriguingly, the concentration of potassium phosphate buffer (KPB) showed significant impact on the aggregation state of Trx-SmL during ultrasonic disruption. The proportion of inclusion bodies increased dramatically with the increase of KPB concentration from almost completely soluble in 10 mM KPB to insoluble in 200 mM KPB. Based on this new finding, a novel method for refolding and purification of recombinant Trx-SmL was developed by one-step ultrasonication. The Trx-SmL was firstly precipitated in 200 mM KPB, washed for three times, and subsequently subjected to ultrasonic process in 10 mM KPB where refolding and purification occurred simultaneously. This established method was proved to be a straightforward, economical, and efficient purification approach to facilely obtain recombinant Trx-SmL protein with high purity (> 90%) and activity recovery yield (> 80%) from cell lysates. The application potential of the purified fusion Trx-SmL was further demonstrated by kinetic bioresolution of (±)-trans-3-(4-methoxyphenyl)glycidic acid methyl ester [(±)-MPGM] producing optically pure (−)-MPGM, a key intermediate for diltiazem, with an overall yield of 41.5% and ee of 99%.

粘质沙雷氏菌SRICI-01（Trx-SmL）的新脂肪酶在大肠杆菌中成功过表达与硫氧还蛋白（Trx）融合标签。有趣的是，在超声破坏过程中，磷酸钾缓冲液（KPB）的浓度对Trx-SmL的聚集状态有显着影响。随着KPB浓度的增加，包涵体的比例从几乎完全溶于10 mM KPB到不溶于200 mM KPB急剧增加。基于这一新发现，通过一步超声处理开发了一种新的重组Trx-SmL的折叠和纯化方法。Trx-SmL首先在200 mM KPB中沉淀，洗涤3次，然后在10 mM KPB中进行超声处理，在此同时发生重折叠和纯化。这种既定方法被证明是一种简单，经济，高效的纯化方法从细胞裂解物中轻松获得高纯度（> 90％）和活性回收率（> 80％）的重组Trx-SmL蛋白。纯化的融合Trx-SmL的应用潜力通过（±）-生成光学纯的（-）-MPGM（地尔硫卓的关键中间体）的反式-3-（4-甲氧基苯基）缩水甘油酸甲酯[（±）-MPGM]，总收率为41.5％，ee为99％。