Shen Yanfei, Wang Fanghua, Lan Dongming, Liu Yuanyuan, Yang Bo, Wang Yonghua
Department of Biotechnology, School of Bioscience & Bioengineering, South China University of Technology, Guangzhou 510006, Guangdong, China; E-Mail:
Int J Mol Sci. 2011;12(11):7609-25. doi: 10.3390/ijms12117609. Epub 2011 Nov 7.
Experiments were carried out to investigate the effects of various factors on the activity and conformation of recombinant leucine aminopeptidase of Bacillus kaustophilus CCRC 11223 (BkLAP) and potential utilization of BkLAP in the hydrolysis of anchovy protein. Optimal temperature and pH of BkLAP were 70 °C and 8.0 in potassium-phosphate buffer, respectively, and the activity was strongly stimulated by Ni(2+), followed by Mn(2+) and Co(2+). Conformational studies via circular dichroism spectroscopy indicated that various factors could influence the secondary structure of BkLAP to different extents and further induce the changes in enzymatic activity. The secondary structure of BkLAP was slightly modified by Ni(2+) at the concentration of 1×10(-4) M, however, significant changes on the secondary structures of the enzyme were observed when Hg(2+) was added to the concentration of 1×10(-4) M. The potential application of BkLAP was evaluated through combination with the commercial or endogenous enzyme to hydrolysis the anchovy protein. Results showed that combining the BkLAP with other enzymes could significantly increase the degree of hydrolysis and amino acid component of hydrolysate. In this regard, BkLAP is a potential enzyme that can be used in the protein hydrolysate industry.
开展了实验以研究各种因素对嗜碱芽孢杆菌CCRC 11223重组亮氨酸氨肽酶（BkLAP）活性和构象的影响，以及BkLAP在水解鳀鱼蛋白方面的潜在应用。在磷酸钾缓冲液中，BkLAP的最佳温度和pH分别为70℃和8.0，其活性受到Ni(2+)的强烈刺激，其次是Mn(2+)和Co(2+)。通过圆二色光谱进行的构象研究表明，各种因素可在不同程度上影响BkLAP的二级结构，并进一步引起酶活性的变化。当Ni(2+)浓度为1×10(-4) M时，BkLAP的二级结构略有改变，然而，当加入浓度为1×10(-4) M的Hg(2+)时，观察到该酶二级结构有显著变化。通过将BkLAP与商业酶或内源酶结合水解鳀鱼蛋白来评估其潜在应用。结果表明，将BkLAP与其他酶结合可显著提高水解度和水解产物的氨基酸组成。就此而言，BkLAP是一种可用于蛋白水解物行业的潜在酶。
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