The reduction of nitrite to nitric oxide (NO) in denitrifying bacteria is catalysed by the enzyme nitrite reductase, which, in several species, is a haem-containing protein with one c haem and one d1 haem per monomer (cd1NIR), encoded by the nirS gene. The structure and function of cd1NIR is reviewed here, together with the biogenesis of its cofactors. Mechanistic studies of the catalytic cycle have revealed that the high affinity for nitrite, the allosteric control of electron transfer and the fast NO dissociation are distinctive features of cd1NIR. It is clear that the peculiar d1 haem cofactor, unique to cd1NIRs, has evolved to have low affinity for NO as compared to other haem, thus allowing the enzyme to cope with NO safely during turnover when both nitrite and electrons are available. cd1NIR is a potential tool for engineering biosensors and bioreactors for green technology applications. On the other hand, the nirS gene represents a functional biomarker for investigating the phylogenetic diversity of bacterial communities of denitrifiers and anaerobic ammonium oxidation bacteria. cd1NIR also plays a relevant role in bacterial pathogens, where its expression is socially regulated in order to control pathogenicity and community behaviours, such as biofilm formation in chronic infections.