Structural basis of microcystinase activity for biodegrading microcystin-LR. (December 2019)

• Record Type: Journal Article
• Authors: Xu, Qianqian; Fan, Jinhui; Yan, Hai; Ahmad, Shahbaz; Zhao, Zhenzhen; Yin, Chunhua; Liu, Xiaolu; Liu, Yang; Zhang, Haiyang
• Is Part Of: Chemosphere. Volume 236(2019)
• Publication Date: 2019-12
• Subjects: Microcystin biodegradation -- Microcystinase -- Molecular simulation -- Site-directed mutagenesis -- Enzymatic mechanism; Pollution -- Periodicals; Pollution -- Physiological effect -- Periodicals; Environmental sciences -- Periodicals; Atmospheric chemistry -- Periodicals; 551.511
• DOI: 10.1016/j.chemosphere.2019.07.012
• Languages: English
• ISSNs: 0045-6535

Abstract

Abstract: Microcystinase (MlrA) catalyzes the first and most important biodegradation step of hepatotoxic microcystin-LR (MC-LR) produced and released from cyanobacterial cells, and the underlying catalytic mechanism is not completely understood yet. MlrA was postulated previously to be a metalloprotease with an active site of H 260 AIH 263 NE 265, a variant of the common metal-binding motif of HEXXH. Through comparison with representative modes in HEXXH-containing metalloproteases, molecular dynamics simulation, homology modeling, and docking, the active sites of MlrA involved in the MC-LR biodegradation by Sphingomonas sp . USTB-05 were predicted. Site-directed mutants of MlrA were constructed for verification then. The results show that MlrA is likely not a metalloprotease, but a glutamate protease belonging to type II CAAX prenyl endopeptidases. Combined with the biodegradation of MC-LR by MlrA and its mutants, a complete enzymatic mechanism for MC-LR biodegradation by MlrA is proposed: Glu 172 and His 205 activate a water molecule facilitating a nucleophilic attack on the Adda-Arg peptide bond of MC-LR; Trp 176 and Trp 201 contact the carboxylate side chain of Glu 172 and, by raising its p K a potentially, accelerate the reaction rates; His 260 and Asn 264 (located in the previous postulated active center of H 260 AIH 263 NE 265 ) function as an oxyanion hole to stabilize the transition states. This study reveals the enzymatic mechanism of MlrA for catalyzing MC-LR in both the representative modes and the experiments of site-directed mutagenesis. Graphical abstract: Image 1 Highlights: Microcystinase (MlrA) was predicted belonging to type II CAAX prenyl endopeptidases. E172, H205, H260, and N264 are critical residues responsible for MlrA activity. Homology modeling of MlrA was verified positively by site-directed mutagenesis.