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A “bridge-like” structure responsible for the substrate selectivity of mono- and diacylglycerol lipase from Aspergillus oryzae

Liu, Lu, Lan, Dongming, Wang, Qian, Gao, Chongliang, Li, Zhigang, Yang, Bo, Wang, Yonghua
Journal of Molecular Catalysis. B, Enzymatic 2013 v.97 pp. 144-149
Aspergillus oryzae, active sites, catalytic activity, consensus sequence, enzyme substrates, lipoprotein lipase, molecular dynamics, site-directed mutagenesis, triacylglycerol lipase, triacylglycerols
Lipase from Aspergillus oryzae (AOL) was experimentally identified to be strictly specific for mono- and diacylglycerol, but not triacylglycerol, whose substrate selectivity was considerably different from most of lipases. The mechanism of this unique feature was still not clear. To address this problem, the structure of AOL in complex with substrate analog was constructed and molecular dynamics simulations were performed to fully relax steric clashes occurring in this enzyme-substrate complex. The bound diacylglycerol analog unambiguously identified two binding pockets, into which the corresponding parts of substrate were closely fitted. Comparison of AOL with the triacylglycerol lipase in homology revealed that its substrate selectivity mainly came from its unique binding pocket: a “bridge-like” structure was created by two bulky residues (W89 and F257) on the top of catalytic site, which prevented substrates with large size from entering the active site. In addition, a common feature that was responsible for substrate selectivity of mono- and diacylglycerol lipases (MDGL) was drawn. A well-conserved sequence motif F-X-X-H was identified, and this consensus sequence participated in substrate recognition of these MDGLs by forming the “bridge-like” structure that governed the specific binding of substrate. These results could provide a guide for discovering novel MDGLs and pave the way for site-directed mutagenesis experiments to improve enzyme for applications.