The Molecular Structure and Catalytic Mechanism of a Quorum-Quenching N-Acyl-L-Homoserine Lactone Hydrolase

In many Gram-negative bacteria, including a number of pathogens such as Pseudomonas aeruginosa and Erwinia carotovora, virulence factor production and biofilm formation are linked to the quorum-sensing systems that use diffusible N-acyl-L-homoserine lactones (AHLs) as intercellular messenger molecul...

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Published inProceedings of the National Academy of Sciences - PNAS Vol. 102; no. 49; pp. 17606 - 17611
Main Authors Myung Hee Kim, Won-Chan Choi, Hye Ok Kang, Jong Suk Lee, Beom Sik Kang, Kyung-Jin Kim, Zygmunt S. Derewenda, Tae-Kwang Oh, Choong Hwan Lee, Lee, Jung-Kee
Format Journal Article
LanguageEnglish
Published United States National Academy of Sciences 06.12.2005
National Acad Sciences
Subjects
Atoms
Bacillus thuringiensis
Bacillus thuringiensis - enzymology
Bacillus thuringiensis - genetics
Bacteria
Binding Sites
Biochemistry
Biofilms
Biological Sciences
Carboxylic Ester Hydrolases - chemistry
Carboxylic Ester Hydrolases - genetics
Carboxylic Ester Hydrolases - metabolism
Catalysis
Cations, Divalent - chemistry
Crystal structure
Enzyme substrates
Enzymes
Erwinia carotovora
Hydrogen bonds
Ions
Lactones
Models, Molecular
Molecules
Mutation - genetics
Oxygen
Protein Structure, Tertiary
Pseudomonas aeruginosa
Substrate Specificity
Zinc
Zinc - chemistry
Zinc - metabolism
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Summary:In many Gram-negative bacteria, including a number of pathogens such as Pseudomonas aeruginosa and Erwinia carotovora, virulence factor production and biofilm formation are linked to the quorum-sensing systems that use diffusible N-acyl-L-homoserine lactones (AHLs) as intercellular messenger molecules. A number of organisms also contain genes coding for lactonases that hydrolyze AHLs into inactive products, thereby blocking the quorum-sensing systems. Consequently, these enzymes attract intense interest for the development of antiinfection therapies. However, the catalytic mechanism of AHL-lactonase is poorly understood and subject to controversy. We here report a 2.0-Å resolution structure of the AHL-lactonase from Bacillus thuringiensis and a 1.7-Å crystal structure of its complex with L-homoserine lactone. Despite limited sequence similarity, the enzyme shows remarkable structural similarities to glyoxalase II and RNase Z proteins, members of the metallo-β-lactamase superfamily. We present experimental evidence that AHL-lactonase is a metalloenzyme containing two zinc ions involved in catalysis, and we propose a catalytic mechanism for bacterial metallo-AHL-lactonases.
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Edited by Harry B. Gray, California Institute of Technology, Pasadena, CA, and approved October 13, 2005
Conflict of interest statement: No conflicts declared.
This paper was submitted directly (Track II) to the PNAS office.
Author contributions: M.H.K., T.-K.O., C.H.L., and J.-K.L. designed research; M.H.K., W.-C.C., H.O.K., J.S.L., B.S.K., and K.-J.K. performed research; M.H.K., W.-C.C., and J.S.L. contributed new reagents/analytic tools; M.H.K., W.-C.C., H.O.K., J.S.L., B.S.K., K.-J.K., Z.S.D., T.-K.O., C.H.L., and J.-K.L. analyzed data; and M.H.K., Z.S.D., C.H.L., and J.-K.L. wrote the paper.
To whom correspondence may be addressed. E-mail: chlee@kribb.re.kr or jklee@kribb.re.kr.
Data deposition: The atomic coordinates and structure factors have been deposited in the Protein Data Bank, www.pdb.org (PDB ID codes 2BTN and 2BR6).
Abbreviations: QS, quorum sensing; AHL, N-acyl-l-homoserine lactone; HSL, homoserine lactone; BTK-AiiA, AHL-lactonase from Bacillus thuringiensis subsp. kurstaki HD263; C6-AHL, N-hexanoyl-l-homoserine lactone; LC-ESI-MS, liquid chromatography electrospray ionization MS; PDB, Protein Data Bank.
Note. When this paper was at the stage of revision, a similar study was published by Liu et al. (32). This work also described the crystal structure of an N-acyl-l-homoserine lactone hydrolase from B. thuringiensis, albeit only in the apo-form with the active site occupied by a glycerol molecule. There are no discrepancies between the two atomic models, and the conclusions regarding the enzymatic mechanism are similar. However, the structure of the complex with an inhibitory HSL molecule reported in our study allows for a more confident description of the structural roots of substrate specificity and reaction mechanism.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.0504996102