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Structural basis for ligand discrimination and response initiation in the heme-based oxygen sensor FixL
K R Rodgers1, G S Lukat-Rodgers, J A Barron
1Department of Chemistry, North Dakota State University, Fargo 58105, USA.
Biochemistry
|July 23, 1996
Summary
Rhizobium meliloti FixL, a bacterial O2-sensing protein, uses heme ligation to control kinase activity. Spectroscopic and kinetic studies reveal heme pocket structural constraints influencing ligand binding, suggesting a feedback mechanism for O2 sensing.
Area of Science:
- Biochemistry
- Molecular Biology
- Protein Chemistry
Background:
- FixL is a bacterial O2-sensing protein regulating kinase activity via heme ligation.
- Heme spin state transitions modulate FixL kinase activity, but the mechanism remains unclear.
Purpose of the Study:
- Investigate the structural and functional properties of Rhizobium meliloti FixL domains.
- Elucidate the mechanism of O2-dependent kinase inhibition and ligand selectivity.
Main Methods:
- Resonance Raman spectroscopy to characterize heme environments.
- 1H NMR spectroscopy to probe heme pocket structure.
- Kinetic and thermodynamic assays for ligand binding.
Main Results:
- FixL heme pocket is smaller and asymmetric compared to myoglobin.
- Kinase domain imposes steric constraints, affecting ligand binding kinetics and thermodynamics.
- FixL exhibits unique selectivity for small ligands like O2 and CO.
Conclusions:
- The heme pocket's structure, influenced by the kinase domain, dictates ligand selectivity.
- FixL's ligand binding properties are crucial for its role in O2 sensing.
- A mechanism for phosphate-dependent feedback control of heme ligand affinity is proposed.