Structural and Functional Mechanisms of Adaptations of WrbA in Extremophilic Organisms
Abstract
Properties of FMN-containing NADH:quinone oxidoreductases (WrbA) from the hyperthermophile Archaeoglohus fudgidus and the mesophile Escherichia coli were investigated to gain an understanding of the mechanisms by which proteins in extremophilic organisms adapt to high temperature. A model of the thermophilic enzyme was constructed based on the crystal structure of the mesophilic counterpart to guide experiments. An electrochemical cell was designed and constructed to probe the redox properties of FMN. Only subtle differences in the midpoint potential were recorded for the two enzymes consistent with congruent physiological functions. Contrary to redox potentials, the two proteins showed differences in FMN binding. Strong cooperativity in flavin binding for the mesophilic enzyme was dependent on the concentration of FMN whereas binding for the thermophilic enzyme was independent of ligand concentration. Comparison of enzyme activities between the two proteins with a variety of quinones suggested differences in the active site architecture accommodating electron acceptors. Properties of the mesophilic wild-type versus single amino acid substitution variants identified residues adjacent to FMN influencing the redox potential, oligomerization and enzyme activity. Both enzymes were suitable catalysts for biofuel cells and NADH-dependent amperometric NADH biosensors, although the thermophilic enzyme was found to be superior.
Document Details
- Document Type
- Technical Report
- Publication Date
- May 11, 2010
- Accession Number
- ADA567242
Entities
People
- James G. Ferry
Organizations
- Pennsylvania State University