Molecular interplay of an assembly machinery for nitrous oxide reductase.

Emissions of the critical ozone-depleting and greenhouse gas nitrous oxide (NO) from soils and industrial processes have increased considerably over the last decades. As the final step of bacterial denitrification, NO is reduced to chemically inert N (refs. ) in a reaction that is catalysed by the copper-dependent nitrous oxide reductase (NOR) (ref.

The Copper Chaperone NosL Forms a Heterometal Site for Cu Delivery to Nitrous Oxide Reductase.

The final step of denitrification is the reduction of nitrous oxide (N O) to N , mediated by Cu-dependent nitrous oxide reductase (N OR). Its metal centers, Cu and Cu , are assembled through sequential provision of twelve Cu ions by a metallochaperone that forms part of a nos gene cluster encoding the enzyme and its accessory factors. The chaperone is the nosL gene product, an 18 kDa lipoprotein predicted to reside in the outer membrane of Gram-negative bacteria.

Functional assembly of nitrous oxide reductase provides insights into copper site maturation.

The multicopper enzyme nitrous oxide reductase reduces the greenhouse gas NO to uncritical N as the final step of bacterial denitrification. Its two metal centers require an elaborate assembly machinery that so far has precluded heterologous production as a prerequisite for bioremediatory applications in agriculture and wastewater treatment. Here, we report on the production of active holoenzyme in using a two-plasmid system to produce the entire biosynthetic machinery as well as the structural gene for the enzyme.