Title
Consensus design for improved thermostability of lipoxygenase from Anabaena sp. PCC 7120
Author
Hui Qian, Chong Zhang, Zhaoxin Lu, Bingjie Xia, Xiaomei Bie, Haizhen Zhao, Fengxia Lu, Guang-Yu Yang
Year
2018
Journal
BMC Biotechnology
Abstract
Lipoxygenase (LOX) from Anabaena sp. PCC 7120 (Ana-rLOX) offers important applications in the food industry, especially for improving aroma and dough rheological properties. However, industrial applications of LOXs have been limited by their poor thermostability. Herein, we report a bioinformatics-based consensus concept approach for the engineering of thermostable Ana-rLOX. A series of mutations (N130D, G260A, S437T, N130D/G260Q, N130D/S437Y) showed higher thermostability and activity than the wild-type enzyme. Thus, N130D/G260Q exhibited a 6.6-fold increase in half-life and 2.45 °C increase in unfolding temperature; N130D/S437Y showed a 10 °C increase in optimal temperature. The secondary structure did not change much that contributed to improved thermostability were investigated in detail using circular dichroism. Homology modeling suggested that enhanced thermostability and specific activity may result from favorable hydrophobic interactions. A series of mutations were achieved, showing higher thermostability and activity than the wild-type enzyme by semi-rational mutagenesis with limited structure information. Our findings provide important new insights into molecular modifications aimed at improving Ana-rLOX thermostability and activity.
Instrument
J-810
Keywords
Circular dichroism, Secondary structure, Biochemistry