Catalysis and oxygen binding of Ec DOS: a haem-based oxygen-sensor enzyme from Escherichia coli
| Autor: | Atsunari Tanaka, Toru Shimizu, Jotaro Igarashi, Yukako Ishitsuka, Hiroto Takahashi, Kazuo Kobayashi, Nao Yokota |
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| Rok vydání: | 2010 |
| Předmět: |
Stereochemistry
Dimer Allosteric regulation Kinetics Heme Plasma protein binding Ligands Biochemistry chemistry.chemical_compound Bacterial Proteins Catalytic Domain Escherichia coli Molecule Cloning Molecular Molecular Biology Phosphoric Diester Hydrolases Chemistry Ligand digestive oral and skin physiology General Medicine Protein Structure Tertiary Oxygen Protein Subunits Crystallography Biocatalysis Mutagenesis Site-Directed Protein Multimerization Pulse Radiolysis Oxidation-Reduction Oxygen binding Protein Binding |
| Zdroj: | The Journal of Biochemistry. 148:693-703 |
| ISSN: | 1756-2651 0021-924X |
| Popis: | A phosphodiesterase (PDE) from Escherichia coli (Ec DOS) is a novel haem-based oxygen sensor enzyme. Binding of O(2) to the reduced haem in the sensor domain enhances PDE activity exerted by the catalytic domain. Kinetic analysis of oxygen-dependent catalytic enhancement showed a sigmoidal curve with a Hill coefficient value of 2.8. To establish the molecular mechanism underlying allosteric regulation, we analysed binding of the O(2) ligand following reduction of haem in the isolated dimeric sensor domain using pulse radiolysis. Spectral changes accompanying O(2) binding were composed of two phases as a result of reduction of two haem complexes when high-dose electron beams were applied. In contrast, upon reduction of the dimer with a low-dose beam, the kinetics of O(2) ligation displayed single-phase behaviour as a result of the reduction of one haem complex within dimer. Based on these results, we propose that the faster phase corresponds to binding of the first O(2) molecule to one subunit of the dimer, followed by binding of the second O(2) molecule to the other subunit. Notably, for the haem axial ligand mutant proteins, M95A and M95L, O(2) binding displayed single-phase kinetics and was independent of electron beam dose. |
| Databáze: | OpenAIRE |
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