Enlarging the gas access channel to the active site renders the regulatory hydrogenase HupUV of Rhodobacter capsulatus O2 sensitive without affecting its transductory activity.

摘要: In the photosynthetic bacterium Rhodobacter capsulatus, synthesis of energy-producing hydrogenase, HupSL, is regulated by substrate H2, which detected a regulatory HupUV. The HupUV protein exhibits typical features [NiFe] hydrogenases but, interestingly, resistant to inactivation O2. Understanding O2 resistance will help in design with high potential for biotechnological applications. To test whether this property results from inaccessibility active site, we introduced two mutations order enlarge gas access channel protein. We showed that such (Ile65Val and Phe113Leu HupV) rendered sensitive inactivation. Also, contrast wild-type protein, mutated exhibited an increase hydrogenase activity after reductive activation presence reduced methyl viologen (up 30% wild-type). H2-sensing first component H2-transduction cascade, which, together two-component system HupT/HupR, regulates HupSL response H2 availability. vitro, purified mutant was able interact histidine kinase HupT. vivo, same as enzyme equally repress absence H2.