Kinetic, spectral, and structural studies of the slow-binding inhibition of the Escherichia coli dihydrodipicolinate synthase by 2, 4-oxo-pentanoic acid
| Autor: | William E. Karsten, Christian Fleming, Leonard M. Thomas, Priscilla Seabourn, Lilian Chooback, Christina G. Bruxvoort |
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| Rok vydání: | 2021 |
| Předmět: |
0301 basic medicine
Dihydrodipicolinate synthase Stereochemistry Lysine Biophysics Crystallography X-Ray Biochemistry Article Enamine 03 medical and health sciences chemistry.chemical_compound Reaction rate constant Catalytic Domain Escherichia coli Enzyme Inhibitors Pyruvates Molecular Biology Hydro-Lyases 030102 biochemistry & molecular biology biology Chemistry Spectrum Analysis Substrate (chemistry) Hydrogen Bonding Rate-determining step Enzyme structure Molecular Docking Simulation Kinetics 030104 developmental biology Enzyme inhibitor biology.protein Protein Binding |
| Zdroj: | Arch Biochem Biophys |
| ISSN: | 0003-9861 |
| DOI: | 10.1016/j.abb.2021.108819 |
| Popis: | Dihydrodipicolinate synthase (DHDPS) catalyzes the first step in the biosynthetic pathway for production of l -lysine in bacteria and plants. The enzyme has received interest as a potential drug target owing to the absence of the enzyme in mammals. The DHDPS reaction is the rate limiting step in lysine biosynthesis and involves the condensation of l -aspartate-β-semialdehyde and pyruvate to form 2, 3-dihydrodipicolinate. 2, 4-oxo-pentanoic acid (acetopyruvate) is a slow-binding inhibitor of DHDPS that is competitive versus pyruvate with an initial Ki of about 20 μM and a final inhibition constant of about 1.4 μM. The enzyme:acetopyruvate complex displays an absorbance spectrum with a λmax at 304 nm and a longer wavelength shoulder. The rate constant for formation of the complex is 86 M−1 s−1. The enzyme forms a covalent enamine complex with the first substrate pyruvate and can be observed spectrally with a λmax at 271 nm. The spectra of the enzyme in the presence of pyruvate and acetopyruvate shows the initial formation of the pyruvate enamine intermediate followed by the slower appearance of the E:acetopyruvate spectra with a rate constant of about 0.013 s−1. The spectral studies suggest the formation of a Schiff base between acetopyruvate and K161 on enzyme that subsequently deprotonates to form a resonance stabilized anion similar to the enamine intermediate formed with pyruvate. The crystal structure of the E:acetopyruvate complex confirms the formation of the Schiff base between acetopyruvate and K161. |
| Databáze: | OpenAIRE |
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