Immunological and molecular evidence for a membrane-bound, dimeric hydrogenase in Rhodopseudomonas capsulata

Abstract

Hydrogenase from Rhodopseudomonas capsulata membranes was purified to a specific activity of 37 μmol H 2 oxidized·min −1·mg −1. Analysis of the purified hydrogenase by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed major protein-staining bands at 67 and 31 kDa that comigrated with the large and small subunits of Azotobacter vinelandii hydrogenase. Western immunoblots and enzyme-linked immunosorbent assays in microtiter plates demonstrated that the 67 and 31 kDa proteins in the R. capsulata hydrogenase are immunologically related to the large and small subunits, respectively, of the dimeric hydrogenases purified from A. vinelandii, Rhizobium japonicum, Alcaligenes latus, and Alcaligenes eutrophus H16 (membrane-bound). It is concluded that the photosynthetic microorganism, R. capsulata, contains a membrane-bound, dimeric hydrogenase that is immunologically related to the membrane-bound hydrogenase from the aerobic microorganisms mentioned.