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J. Biol. Chem., Vol. 283, Issue 9, 5370-5379, February 29, 2008

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Structural Basis for Catalytic and Inhibitory Mechanisms of β-Hydroxyacyl-acyl Carrier Protein Dehydratase (FabZ)^*

Liang Zhang¹, Weizhi Liu¹, Tiancen Hu, Li Du, Cheng Luo, Kaixian Chen, Xu Shen², and Hualiang Jiang³

From the Drug Discovery and Design Center, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 201203, China and the School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China

β-Hydroxyacyl-acyl carrier protein dehydratase (FabZ) is an important enzyme for the elongation cycles of both saturated and unsaturated fatty acids biosyntheses in the type II fatty acid biosynthesis system (FAS II) pathway. FabZ has been an essential target for the discovery of compounds effective against pathogenic microbes. In this work, to characterize the catalytic and inhibitory mechanisms of FabZ, the crystal structures of the FabZ of Helicobacter pylori (HpFabZ) and its complexes with two newly discovered inhibitors have been solved. Different from the structures of other bacterial FabZs, HpFabZ contains an extra short two-turn -helix (4) between 3 and β3, which plays an important role in shaping the substrate-binding tunnel. Residue Tyr-100 at the entrance of the tunnel adopts either an open or closed conformation in the crystal structure. The crystal structural characterization, the binding affinity determination, and the enzymatic activity assay of the HpFabZ mutant (Y100A) confirm the importance of Tyr-100 in catalytic activity and substrate binding. Residue Phe-83 at the exit tunnel was also refined in two alternative conformations, leading the tunnel to form an L-shape and U-shape. All these data thus contributed much to understanding the catalytic mechanism of HpFabZ. In addition, the co-crystal structures of HpFabZ with its inhibitors have suggested that the enzymatic activity of HpFabZ could be inhibited either by occupying the entrance of the tunnel or plugging the tunnel to prevent the substrate from accessing the active site. Our study has provided some insights into the catalytic and inhibitory mechanisms of FabZ, thus facilitating antibacterial agent development.

Received for publication, July 6, 2007 , and in revised form, December 18, 2007.

^* This work was supported in part by the State Key Program of Basic Research of China (Grants 2004CB58905, 2007CB914304, and 2006AA09Z447), the National Natural Science Foundation of China (Grants 30525024, 90713046, 20721003), Shanghai Basic Research Project (Grants 06JC14080, 03DZ19228) and Foundation of Chinese Academy of Sciences (Grant KSCX1-YW-R-18). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The atomic coordinates and structure factors (code 2GLL, 2GLP, 2GLM, and 2GLV) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. S1–S4 and experimental data.

¹ Both authors contributed equally to this work.

² To whom correspondence may be addressed. E-mail: xshen{at}mail.shcnc.ac.cn. ³ To whom correspondence may be addressed. Tel.: 86-21-50805873; Fax: 86-21-50807088; E-mail: hljiang{at}mail.shcnc.ac.cn.

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