Partial purification and characterization of two hydrogenases from the extreme thermophile Methanococcus jannaschii

Abstract

F420-nonreactive and F420-reactive hydrogenases have been partially purified from Methanococcus jannaschii, an extremely thermophilic methanogen isolated from a submarine hydrothermal vent. The molecular weights of both hydrogenases were determined by native gradient electrophoresis in 5 to 27% polyacrylamide gels. The F420-nonreactive hydrogenase produced one major band (475 kilodaltons), whereas the F420-reactive hydrogenase produced two major bands (990 and 115 kilodaltons). The F420-nonreactive hydrogenase consisted of two subunits (43 and 31 kilodaltons), and the F420-reactive hydrogenase contained three subunits (48, 32, and 25 kilodaltons). Each hydrogenase was active at very high temperatures. Methyl viologen-reducing activity of the F420-nonreactive hydrogenase was maximal at 80°C but was still detectable at 103°C. The maximum activities of F420-reactive hydrogenase for F420 and methyl viologen were measured at 80 and 90°C, respectively. Low but measureable activity toward methyl viologen was repeatedly observed at 103°C. Moreover, the half-life of the F420-nonreactive hydrogenase at 70°C was over 9 h, and that of the F420-reactive enzyme was over 3 h.