The purple (Sulphur) phototrophic bacterium, Thiocapsa roseopersicina BBS contains several [NiFe] hydrogenases, of which two are membrane bound. Mutant T. roseopersicina cells, carrying deletions in both gene clusters showed hydrogenase activity. This activity was located in the cytoplasm. The structural gene cluster hoxEFUYH was identified and sequenced. In addition, genes homologous to hupUV/hoxBC, the hydrogen sensing hydrogenase have been identified and sequenced. Regulation of hydrogenase biosynthesis was studied in detail for HydSL (renamed HynSL). A random mutagenesis system was optimised for T roseopersicina. One of the mutations was in a gene similar to that coding for the HypF proteins in other organisms. Inactivation of the hypF gene resulted in a 60-fold increase in hydrogen evolution under nitrogen fixing conditions. In addition to hypF, the following accessory genes were identified: hydD, hupK, hypC1, hypC2, hypDE. Characterisation of the corresponding gene products and search for additional accessory genes are in progress. (C) 2002 International Association for Hydrogen Energy. Published by Elsevier Science Ltd. All rights reserved.