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Changes in hydrogen-bond strengths explain reduction potentials in 10 rubredoxin variants
I-Jin Lin1, Erika B Gebel, Timothy E Machonkin
1Graduate Program in Biophysics, National Magnetic Resonance Facility at Madison, University of Wisconsin, 433 Babcock Drive, Madison, WI 53706, USA.
Summary
Protein sequence changes alter the reduction potential of iron-sulfur proteins by modifying hydrogen bond strengths. This study quantizes these changes in rubredoxin variants, linking sequence to electrochemical properties.
Area of Science:
- Biochemistry
- Biophysics
- Structural Biology
Background:
- Rubredoxin from Clostridium pasteurianum (CpRd) is a model iron-sulfur protein.
- Protein sequence variations can significantly impact the active site's reduction potential.
Purpose of the Study:
- To investigate how protein sequence modulates the reduction potential in CpRd.
- To correlate hydrogen bond strengths with observed reduction potentials in wild-type and mutant CpRd.
Main Methods:
- Utilized (15)N NMR spectroscopy to measure hydrogen bond strengths.
- Analyzed electron spin delocalization across hydrogen bonds in oxidized (Fe(III)) and reduced (Fe(II)) states.
- Studied wild-type CpRd and nine mutants.
Main Results:
- Quantified strengths of six key hydrogen bonds involving cysteine ligatures.
- Found shorter aggregate hydrogen bond lengths in variants with higher reduction potentials.
- Observed a linear correlation between changes in hydrogen bonding upon reduction and published reduction potentials.
Conclusions:
- Protein sequence effects on CpRd reduction potential are fully explained by alterations in hydrogen bond strengths.
- Hydrogen bond length is a critical determinant of redox potential in iron-sulfur proteins.