Abstract

The membrane-bound hydrogenase of the photosynthetic bacterium Rhodospirillum rubrum has been purified 490-fold with a yield of 5.8%. The enzyme was homogeneous by disc gel electrophoresis. A method for the permanent, oxygen-insensitive, staining of hydrogenase on polyacrylamide gels is described. The enzyme is a monomer of molecular weight about 66,000 containing four iron and four acid-labile sulfur atoms per molecule. The electron paramagnetic resonance spectrum at 20 °K exhibits a strong signal in the oxidized state only with g > 2—this is characteristic of high potential iron-sulfur protein. The hydrogenase is thermostable and also resistant to both denaturation agents and oxygen inactivation. Carbon monoxide reversibly inhibits the enzyme but metal-complexing and thiol-blocking reagents have little effect on activity. The enzyme will catalyze both H 2 evolution and H 2 uptake in the presence of many artificial electron carriers but the two activities differ in their pH optima. There is a correlation between H 2 evolution activity and the redox potential of the mediator dye. Ferredoxins and pyridine nucleotides do not readily interact with the hydrogenase. We have shown that irradiation of a solution containing methyl viologen, EDTA, proflavin, and R. rubrum hydrogenase will evolve hydrogen continuously for over 9 h. However, the enzyme evolves hydrogen at only very low rates from in vitro chloroplast-ferredoxin and chloroplast-methyl viologen systems. R. rubrum hydrogenase has a number of properties in common with the hydrogenases purified from two other photosynthetic bacteria, Chromatium and Thiocapsa, but is distinct from the hydrogenases of nonphotosynthetic bacteria.

Talk to us

Join us for a 30 min session where you can share your feedback and ask us any queries you have

Schedule a call

Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.