Background: Due to its capacity to produce large amounts of cellulases, Trichoderma reesei is increasingly been researched in various fields of white biotechnology, especially in biofuel production from lignocellulosic biomass. The commercial enzyme mixtures produced at industrial scales are not well characterized, and their proteinaceous components are poorly identified and quantified. The development of proteomic methods has made it possible to comprehensively overview the enzymes involved in lignocellulosic biomass degradation which are secreted under various environmental conditions.

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The physiological properties of a hyd mutant of Desulfovibrio vulgaris Hildenborough, lacking periplasmic Fe-only hydrogenase, have been compared with those of the wild-type strain. Fe-only hydrogenase is the main hydrogenase of D. vulgaris Hildenborough, which also has periplasmic NiFe-and NiFeSe-hydrogenases. The hyd mutant grew less well than the wild-type strain in media with sulfate as the electron acceptor and H 2 as the sole electron donor, especially at a high sulfate concentration. Although the hyd mutation had little effect on growth with lactate as the electron donor for sulfate reduction when H 2 was also present, growth in lactateand sulfate-containing media lacking H 2 was less efficient. The hyd mutant produced, transiently, significant amounts of H 2 under these conditions, which were eventually all used for sulfate reduction. Sulfate-reducing bacteria of the genus Desulfovibrio contain the genes for several hydrogenases, including the hynBA and hysBA genes for the NiFe-and NiFeSe-hydrogenases, the hydAB genes for Fe-only hydrogenase, and the hndABCD genes for NADP-reducing hydrogenase. The first three enzymes are translocated by the twin-arginine translocation (tat) system and are thus either periplasmic or membrane bound with the active site facing the periplasm. Only the NADPreducing hydrogenase has been shown elsewhere to be cytoplasmic (11). Desulfovibrio vulgaris strain Hildenborough has the Fe-only hydrogenase as well as the NiFe-and NiFeSehydrogenases. The former is a soluble periplasmic enzyme, whereas the latter two are membrane bound. Searching of the database for the D. vulgaris genome at http://www.tigr.org indicated that D. vulgaris has an HndD homolog but lacks the hndA, hndB, and hndC genes. D. vulgaris is thus unlikely to have a cytoplasmic, NADP ϩ -reducing hydrogenase. The hydAB genes encode the 46-kDa ␣ and the 10-kDa ␤ subunit of Fe-only hydrogenase from D. vulgaris (25). The structures of Fe-only hydrogenase have been determined recently both for CpI, the cytoplasmic enzyme from Clostridium pasteurianum (16), and for the periplasmic enzyme from Desulfovibrio desulfuricans (14). The sequences of the ␣ and ␤ subunits of the periplasmic enzyme in D. desulfuricans and D. vulgaris form a contiguous, single polypeptide of 60 kDa in CpI. The splitting of the sequence into two polypeptides in Desulfovibrio spp. is for export: the ␤ subunit has a long twinarginine-type signal sequence for this purpose (24). The function of CpI in the fermentative metabolism of C. pasteurianum is to reoxidize reduced ferredoxin, using protons as the electron acceptor to produce H 2 (2). A similar function has been proposed elsewhere for Fe-only hydrogenase in lactate metabolism by D. vulgaris (23). Reduction of the Fe-only hydrogenase content by expression of hydAB antisense RNA reduced the growth rate and growth yield of D. vulgaris in lactate-and sulfate-containing medium. Observation of a reduced H 2 burst in the initial stages of growth on this medium also pointed to a decreased H 2 production act...