Hydrogenases (H 2 ases) are metalloproteins. The great majority of them contain iron-sulfur clusters and two metal atoms at their active center, either a Ni and an Fe atom, the [NiFe]-H 2 ases, or two Fe atoms, the [FeFe]-H 2 ases. Enzymes of these two classes catalyze the reversible oxidation of hydrogen gas (H 2<--> 2 H++ 2 e-) and play a central role in microbial energy metabolism; in addition to their role in fermentation and H 2 respiration, H 2 ases may interact with membrane-bound electron transport systems in order to maintain redox poise, particularly in some photosynthetic microorganisms such as cyanobacteria. Recent work has revealed that some H 2 ases, by acting as H 2-sensors, participate in the regulation of gene expression and that H 2-evolving H 2 ases, thought to be involved in purely fermentative processes, play a role in membrane-linked energy conservation through the generation of a protonmotive force. The Hmd hydrogenases of some methanogenic archaea constitute a third class of H 2 ases, characterized by the absence of Fe-S cluster and the presence of an iron-containing cofactor with catalytic properties different from those of [NiFe]-and [FeFe]-H 2 ases. In this review, we emphasise recent advances that have greatly increased our knowledge of microbial H 2 ases, their diversity, the structure of their active site, how the metallocenters are synthesized and assembled, how they function, how the synthesis of these enzymes is controlled by external signals, and their potential use in biological H 2 production.