NMR solution structures of two mutants of desulforedoxin
B J Goodfellow1, F Rusnak, I Moura
1Departamento de Química, Universidade de Aveiro, 3810-193 Aveiro, Portugal. brian.goodfellow@dq.ua.pt
Journal of Inorganic Biochemistry
|January 23, 2003
Summary
Altering the spacing of cysteine residues in Desulfovibrio gigas desulforedoxin (Dx) impacts its structure. Modifying Dx
Area of Science:
- Biochemistry
- Structural Biology
- Protein Science
Background:
- The geometry of metal centers in [Fe-4S] proteins like rubredoxin (Rd) and desulforedoxin (Dx) differs.
- These geometric differences are hypothesized to stem from variations in the spacing of the C-terminal cysteine pair.
Purpose of the Study:
- To investigate the role of C-terminal cysteine spacing in determining the structure of Desulfovibrio gigas desulforedoxin (Dx).
- To compare the structural consequences of altering cysteine spacing in Dx to the structure of rubredoxin (Rd).
Main Methods:
- Site-directed mutagenesis was used to create two Dx mutants with modified cysteine spacing.
- Nuclear Magnetic Resonance (NMR) spectroscopy was employed to determine the solution structures of the wild-type and mutant Dx proteins.
- Structural analysis focused on the impact of amino acid insertions on protein dimerization and metal center geometry.
Main Results:
- Mutant 1 (DxM1), with a single glycine insertion, showed disrupted dimer stability, indicating a shift towards a monomeric state.
- Mutant 3 (DxM3), with a two-amino acid insertion (-P-V-), resulted in a monomeric protein.
- The overall fold near the metal center in DxM3 closely resembled that of rubredoxin (Rd).
Conclusions:
- Cysteine spacing significantly influences the quaternary structure (dimerization) and tertiary structure of Desulfovibrio gigas desulforedoxin.
- Altering cysteine spacing can modulate the structural similarity between desulforedoxin and rubredoxin, providing insights into [Fe-4S] protein evolution and function.
