Structural Determinants Defining the Allosteric Inhibition of an Essential Antibiotic Target

Tatiana P Soares da Costa; Sebastien Desbois; Con Dogovski; Michael A Gorman; Natalia E Ketaren; Jason J Paxman; Tanzeela Siddiqui; Leanne M Zammit; Belinda M Abbott; Roy M Robins-Browne; Michael W Parker; Geoffrey B Jameson; Nathan E Hall; Santosh Panjikar; Matthew A Perugini

doi:10.1016/j.str.2016.05.019

Structural Determinants Defining the Allosteric Inhibition of an Essential Antibiotic Target

Structure. 2016 Aug 2;24(8):1282-1291. doi: 10.1016/j.str.2016.05.019. Epub 2016 Jul 14.

Authors

Affiliations

¹ Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Bundoora, VIC 3086, Australia.
² Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Bundoora, VIC 3086, Australia; Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, VIC 3010, Australia.
³ St Vincent's Institute of Medical Research, Fitzroy, VIC 3065, Australia.
⁴ Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, VIC 3010, Australia.
⁵ Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Bundoora, VIC 3086, Australia; Australian Synchrotron, Clayton, VIC 3168, Australia.
⁶ Department of Chemistry and Physics, La Trobe Institute for Molecular Science, La Trobe University, Bundoora, VIC 3086, Australia.
⁷ Department of Microbiology and Immunology, The University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Parkville, VIC 3010, Australia; Murdoch Childrens Research Institute, Royal Children's Hospital, Parkville, VIC 3052, Australia.
⁸ Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, VIC 3010, Australia; St Vincent's Institute of Medical Research, Fitzroy, VIC 3065, Australia.
⁹ Centre for Structural Biology, Institute of Fundamental Sciences, Massey University, Palmerston North 4442, New Zealand.
¹⁰ Australian Synchrotron, Clayton, VIC 3168, Australia; Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC 3800, Australia.
¹¹ Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Bundoora, VIC 3086, Australia. Electronic address: m.perugini@latrobe.edu.au.

PMID: 27427481
DOI: 10.1016/j.str.2016.05.019

Abstract

Dihydrodipicolinate synthase (DHDPS) catalyzes the first committed step in the lysine biosynthesis pathway of bacteria. The pathway can be regulated by feedback inhibition of DHDPS through the allosteric binding of the end product, lysine. The current dogma states that DHDPS from Gram-negative bacteria are inhibited by lysine but orthologs from Gram-positive species are not. The 1.65-Å resolution structure of the Gram-negative Legionella pneumophila DHDPS and the 1.88-Å resolution structure of the Gram-positive Streptococcus pneumoniae DHDPS bound to lysine, together with comprehensive functional analyses, show that this dogma is incorrect. We subsequently employed our crystallographic data with bioinformatics, mutagenesis, enzyme kinetics, and microscale thermophoresis to reveal that lysine-mediated inhibition is not defined by Gram staining, but by the presence of a His or Glu at position 56 (Escherichia coli numbering). This study has unveiled the molecular determinants defining lysine-mediated allosteric inhibition of bacterial DHDPS.

MeSH terms

Allosteric Regulation
Allosteric Site
Amino Acid Sequence
Binding Sites
Cloning, Molecular
Crystallography, X-Ray
Escherichia coli / enzymology*
Escherichia coli / genetics
Feedback, Physiological*
Gene Expression
Hydro-Lyases / chemistry*
Hydro-Lyases / genetics
Hydro-Lyases / metabolism
Kinetics
Legionella pneumophila / enzymology*
Legionella pneumophila / genetics
Lysine / chemistry*
Lysine / metabolism
Models, Molecular
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Interaction Domains and Motifs
Recombinant Proteins / chemistry
Recombinant Proteins / genetics
Recombinant Proteins / metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Streptococcus pneumoniae / enzymology*
Streptococcus pneumoniae / genetics
Substrate Specificity

Substances

Recombinant Proteins
Hydro-Lyases
4-hydroxy-tetrahydrodipicolinate synthase
Lysine