The efficiency of hydrogen gas production by nitrogenase in bacteria has been improved by the inhibition of antagonistic activity by the uptake hydrogenase. In this study, a mutant lacking the gene coding for the uptake hydrogenase was generated from the photosynthetic beta-proteobacterium Rubrivivax gelatinosus IL144 to explore new ways of hydrogen gas production driven by light energy. The mutant cells produced 25–30% higher amounts of molecular hydrogen than the wild-type cells under nitrogen-deficient conditions under light. Furthermore, by the addition of 5 mM glutamate, the photosynthetic growth rate was greatly enhanced, and the hydrogen gas production activity reached 41.1 (mmol/l) in the mutant.