US EPA

7447003 

Journal Article 

Structural basis of microcystinase activity for biodegrading microcystin-LR 

Xu, Q; Fan, J; Yan, H; Ahmad, S; Zhao, Z; Yin, C; Liu, X; Liu, Y; Zhang, H 

2019 

Yes 

Chemosphere
ISSN: 0045-6535
EISSN: 1879-1298 

Elsevier Ltd 

236 

124281 

English 

31310980 

10.1016/j.chemosphere.2019.07.012 

WOS:000491634500090 

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 H260AIH263NE265, 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: Glu172 and His205 activate a water molecule facilitating a nucleophilic attack on the Adda-Arg peptide bond of MC-LR; Trp176 and Trp201 contact the carboxylate side chain of Glu172and, by raising its pKa potentially, accelerate the reaction rates; His260 and Asn264 (located in the previous postulated active center of H260AIH263NE265) 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. 

Enzymatic mechanism; Microcystin biodegradation; Microcystinase; Molecular simulation; Site-directed mutagenesis; Carboxylation; Catalysis; Metals; Molecular dynamics; Molecules; Mutagenesis; Reaction rates; Enzymatic mechanisms; Microcystinase; Microcystins; Molecular simulations; Site directed mutagenesis; Biodegradation; arginine; carboxylic acid; glutamic acid; metalloproteinase; microcystin LR; microcystinase; prenylamine; proteinase; restriction endonuclease; unclassified drug; microcystin; microcystin LR; biodegradation; catalysis; catalyst; enzyme; enzyme activity; molecular analysis; peptide; reaction rate; simulation; toxin; alpha helix; Article; biodegradation; enzyme active site; enzyme activity; enzyme mechanism; enzyme structure; high performance liquid chromatography; hydrogen bond; Methanococcus maripaludis; molecular dynamics; nonhuman; nucleophilicity; nucleotide sequence; Rhizobium; sequence alignment; site directed mutagenesis; Sphingomonas; structure activity relation; bioremediation; chemical structure; chemistry; Cyanobacteria; Sphingomonas sp.; Biodegradation, Environmental; Microcystins; Molecular Structure