The three-dimensional structure of the enzyme dihydrodipicolinate synthase (KEGG entry Rv2753c, EC 4.2.1.52) from Mycobacterium tuberculosis (Mtb-DHDPS) was determined and refined at 2.28 Å (1 Å=0.1 nm) resolution. The asymmetric unit of the crystal contains two tetramers, each of which we propose to be the functional enzyme unit. This is supported by analytical ultracentrifugation studies, which show the enzyme to be tetrameric in solution. The structure of each subunit consists of an N-terminal (β/α)8-barrel followed by a C-terminal α-helical domain. The active site comprises residues from two adjacent subunits, across an interface, and is located at the C-terminal side of the (β/α)8-barrel domain. A comparison with the other known DHDPS structures shows that the overall architecture of the active site is largely conserved, albeit the proton relay motif comprising Tyr143, Thr54 and Tyr117 appears to be disrupted. The kinetic parameters of the enzyme are reported: KMASA=0.43±0.02 mM, KMpyruvate=0.17±0.01 mM and Vmax=4.42±0.08 μmol·s−1·mg−1. Interestingly, the Vmax of Mtb-DHDPS is 6-fold higher than the corresponding value for Escherichia coli DHDPS, and the enzyme is insensitive to feedback inhibition by (S)-lysine. This can be explained by the three-dimensional structure, which shows that the (S)-lysine-binding site is not conserved in Mtb-DHDPS, when compared with DHDPS enzymes that are known to be inhibited by (S)-lysine. A selection of metabolites from the aspartate family of amino acids do not inhibit this enzyme. A comprehensive understanding of the structure and function of this important enzyme from the (S)-lysine biosynthesis pathway may provide the key for the design of new antibiotics to combat tuberculosis.
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Research Article|
March 27 2008
Crystal structure and kinetic study of dihydrodipicolinate synthase from Mycobacterium tuberculosis
Georgia Kefala;
*EMBL Hamburg Outstation, c/o DESY, Notkestrasse 85, D-22603 Hamburg, Germany
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Genevieve L. Evans;
Genevieve L. Evans
2
†School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand
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Michael D. W. Griffin;
Michael D. W. Griffin
‡Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, 30 Flemington Road, University of Melbourne, Melbourne, Victoria 3010, Australia
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Sean R. A. Devenish;
Sean R. A. Devenish
†School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand
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F. Grant Pearce;
F. Grant Pearce
†School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand
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Matthew A. Perugini;
Matthew A. Perugini
‡Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, 30 Flemington Road, University of Melbourne, Melbourne, Victoria 3010, Australia
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Juliet A. Gerrard;
Juliet A. Gerrard
†School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand
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Manfred S. Weiss;
Manfred S. Weiss
3
*EMBL Hamburg Outstation, c/o DESY, Notkestrasse 85, D-22603 Hamburg, Germany
3Correspondence may be addressed to either of these authors (email msweiss@embl-hamburg.de or rdobson@unimelb.edu.au).
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Renwick C. J. Dobson
Renwick C. J. Dobson
3
‡Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, 30 Flemington Road, University of Melbourne, Melbourne, Victoria 3010, Australia
3Correspondence may be addressed to either of these authors (email msweiss@embl-hamburg.de or rdobson@unimelb.edu.au).
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Publisher: Portland Press Ltd
Received:
October 04 2007
Revision Received:
November 27 2007
Accepted:
December 07 2007
Accepted Manuscript online:
December 07 2007
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2008 Biochemical Society
2008
Biochem J (2008) 411 (2): 351–360.
Article history
Received:
October 04 2007
Revision Received:
November 27 2007
Accepted:
December 07 2007
Accepted Manuscript online:
December 07 2007
Citation
Georgia Kefala, Genevieve L. Evans, Michael D. W. Griffin, Sean R. A. Devenish, F. Grant Pearce, Matthew A. Perugini, Juliet A. Gerrard, Manfred S. Weiss, Renwick C. J. Dobson; Crystal structure and kinetic study of dihydrodipicolinate synthase from Mycobacterium tuberculosis. Biochem J 15 April 2008; 411 (2): 351–360. doi: https://doi.org/10.1042/BJ20071360
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