Crystal structure and site-directed mutagenesis analyses of haloalkane dehalogenase linB from sphingobium sp. Strain MI1205

Masahiko Okai, Jun Ohtsuka, Lica Fabiana Imai, Tomoko Mase, Ryota Moriuchi, Masataka Tsuda, Koji Nagata, Yuji Nagata, Masaru Tanokura

Research output: Contribution to journalArticlepeer-review

20 Citations (Scopus)

Abstract

The enzymes LinBUT and LinBMI (LinB from Sphingobium japonicum UT26 and Sphingobium sp. MI1205, respectively) catalyze the hydrolytic dechlorination of β-hexachlorocyclohexane (β-HCH) and yield different producs, 2,3,4,5,6-pentachlorocyclohexanol (PCHL) and 2,3,5,6-tetrachlorocyclohexane-1,4-diol (TCDL), respectively, despite their 98% identity in amino acid sequence. To reveal the structural basis of their different enzymatic properties, we performed site-directed mutagenesis and X-ray crystallographic studies of LinBMI and its seven point mutants. The mutation analysis revealed that the seven amino acid residues uniquely found in LinBMI were categorized into three groups based on the efficiency of the first-step (from β-HCH to PCHL) and second-step (from PCHL to TCDL) conversions. Crystal structure analyses of wild-type LinBMI and its seven point mutants indicated how each mutated residue contributed to the first- and second-step conversions by LinBMI. The dynamics simulation analyses of wild-type LinBMI and LinBUT revealed that the entrance of the substrateaccess tunnel of LinBUT was more flexible than that of LinBMI, which could lead to the different efficiencies of dehalogenationactivity between these dehalogenases.

Original languageEnglish
Pages (from-to)2642-2651
Number of pages10
JournalJournal of Bacteriology
Volume195
Issue number11
DOIs
Publication statusPublished - 2013 Jun

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