This Article Full Text Full Text (PDF) Supplemental material Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Jitsumori, K. Articles by Esaki, N. Search for Related Content PubMed PubMed Citation Articles by Jitsumori, K. Articles by Esaki, N.

 Previous Article  |  Next Article 

Journal of Bacteriology, April 2009, p. 2630-2637, Vol. 191, No. 8
0021-9193/09/$08.00+0     doi:10.1128/JB.01654-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

X-Ray Crystallographic and Mutational Studies of Fluoroacetate Dehalogenase from Burkholderia sp. Strain FA1 ^,

Keiji Jitsumori,^1, Rie Omi,^1,2, Tatsuo Kurihara,^1* Atsushi Kurata,¹ Hisaaki Mihara,¹ Ikuko Miyahara,² Ken Hirotsu,² and Nobuyoshi Esaki^1*

Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan,¹ Department of Chemistry, Graduate School of Science, Osaka City University, Sumiyoshi-ku, Osaka 558-6111, Japan²

Received 21 November 2008/ Accepted 4 February 2009

Fluoroacetate dehalogenase catalyzes the hydrolytic defluorination of fluoroacetate to produce glycolate. The enzyme is unique in that it catalyzes the cleavage of a carbon-fluorine bond of an aliphatic compound: the bond energy of the carbon-fluorine bond is among the highest found in natural products. The enzyme also acts on chloroacetate, although much less efficiently. We here determined the X-ray crystal structure of the enzyme from Burkholderia sp. strain FA1 as the first experimentally determined three-dimensional structure of fluoroacetate dehalogenase. The enzyme belongs to the /β hydrolase superfamily and exists as a homodimer. Each subunit consists of core and cap domains. The catalytic triad, Asp104-His271-Asp128, of which Asp104 serves as the catalytic nucleophile, was found in the core domain at the domain interface. The active site was composed of Phe34, Asp104, Arg105, Arg108, Asp128, His271, and Phe272 of the core domain and Tyr147, His149, Trp150, and Tyr212 of the cap domain. An electron density peak corresponding to a chloride ion was found in the vicinity of the N¹ atom of Trp150 and the N² atom of His149, suggesting that these are the halide ion acceptors. Site-directed replacement of each of the active-site residues, except for Trp150, by Ala caused the total loss of the activity toward fluoroacetate and chloroacetate, whereas the replacement of Trp150 caused the loss of the activity only toward fluoroacetate. An interaction between Trp150 and the fluorine atom is probably an absolute requirement for the reduction of the activation energy for the cleavage of the carbon-fluorine bond.

---

* Corresponding author. Mailing address for Tatsuo Kurihara and Nobuyoshi Esaki: Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan. Phone for Tatsuo Kurihara: 81-774-38-4710. Phone for Nobuyoshi Esaki: 81-774-38-3240. Fax for Tatsuo Kurihara and Nobuyoshi Esaki: 81-774-38-3248. E-mail for Tatsuo Kurihara: kurihara{at}scl.kyoto-u.ac.jp. E-mail for Nobuyoshi Esaki: esakin{at}scl.kyoto-u.ac.jp

Published ahead of print on 13 February 2009.

Supplemental material for this article may be found at http://jb.asm.org/.

These authors contributed equally to this work and should be considered as equivalent first authors.

---

Journal of Bacteriology, April 2009, p. 2630-2637, Vol. 191, No. 8
0021-9193/09/$08.00+0     doi:10.1128/JB.01654-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.