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Crystal structures of E. coli native MenH and two active site mutants
Jodie M Johnston1, Ming Jiang, Zhihong Guo
1Maurice Wilkins Centre and School of Biological Sciences, University of Auckland, Auckland, New Zealand.
Plos One
|May 3, 2013
Summary
The enzyme MenH, crucial for menaquinone biosynthesis, has a conventional oxyanion hole, differing from previous models. Structural analysis reveals insights into its active site for potential selective inhibition.
Area of Science:
- Biochemistry
- Structural Biology
- Enzymology
Background:
- Menaquinone biosynthesis is essential for bacterial respiration.
- The enzyme 2-succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate synthase (MenH) was recently identified in this pathway.
- Previous studies proposed a non-standard oxyanion hole involving Tyr85 and Trp147 in MenH.
Purpose of the Study:
- To determine the crystal structure of E. coli MenH (EcMenH) and active site mutants.
- To elucidate the active site architecture and substrate binding mechanism of MenH.
- To clarify the nature of the oxyanion hole in MenH.
Main Methods:
- X-ray crystallography to solve the structures of EcMenH and its mutants (Tyr85Phe, Arg124Ala).
- Computational docking studies with the MenH substrate and transition state model.
- Analysis of conserved residues and active site geometry.
Main Results:
- The crystal structure of EcMenH reveals an α/β hydrolase fold with a helical lid covering the active site groove.
- Bound sulfate and chloride anions in the active site suggest binding sites for anionic substrate groups.
- Structural and docking data indicate a conventional oxyanion hole involving peptide NH groups, not Tyr85 and Trp147.
Conclusions:
- The oxyanion hole in MenH is of a conventional nature, consistent with S. aureus MenH structure.
- Differences in the active site periphery between E. coli and S. aureus MenH may allow for selective inhibition.
- Structural insights provide a basis for designing targeted inhibitors of bacterial menaquinone biosynthesis.
