Molecular interplay of an assembly machinery for nitrous oxide reductase

Abstract

Emissions of the critical ozone-depleting and greenhouse gas nitrous oxide (N2O) from soils and industrial processes have increased significantly over the last decades 1-3 . As the final step of bacterial denitrification, the critical greenhouse gas N2O is reduced to chemically inert N2 (ref. 1,4) in a reaction catalyzed by the copper-dependent nitrous oxide reductase (N2OR) 5 . The assembly of its unique [4Cu:2S] active site cluster CuZ requires both the ATP-bindingcassette (ABC) complex NosDFY and the membrane-anchored copper chaperone NosL 4,6 .Here we report cryo-electron microscopy structures of Pseudomonas stutzeri NosDFY, and its complexes with NosL and N2OR, respectively. We find that the periplasmic NosD protein features a binding site for a Cu + ion and interacts specifically with NosL in its nucleotide-free state, while its binding to N2OR requires a conformational change triggered by ATP binding.Mutually exclusive complex structures of NosDFY with NosL and N2OR reveal a sequential metal trafficking and assembly pathway for the most complex copper site known to date. In it, NosDFY acts as a mechanical energy transducer rather than a transporter. It links ATP hydrolysis in the cytoplasm to a conformational transition of the NosD subunit in the periplasm, which is required for NosDFY to switch its interaction partner so that copper ions are handed over from the chaperone NosL to the enzyme N2OR.

Cu4S Cluster in “0-Hole” and “1-Hole” States: Geometric and Electronic Structure Variations for the Active CuZ* Site of N2O Reductase

“…The latter is reminiscent of our 1-hole complex, which indeed undergoes a structural rearrangement from a seesaw to a square planar core geometry. Though the protein environment was suggested to be strained, the flipping of some histidine rings as well as a conformational switch of the active site relevant to its function has indeed been observed for these N 2 OR enzymes , and has been suggested for their Cu A site by theoretical calculations as well . Furthermore, a complex network of hydrogen bonds involving the protein environment has been suggested to determine the orientation of the histidine side chains .…”

What can molecular assembly learn from catalysed assembly in living organisms?

“…membrane. 73 Recently, Mu ¨ller et al found that this step is assisted by two membrane proteins NosL and NosDFY (Fig. 6B).…”

Metagenomic insight into antagonistic effects of a nitrification inhibitor and phosphate‐solubilizing bacteria on soil protease activity

“…Soil mineral N is one of the most important nutrients to favor soil microbial growth and improve plant production (Fan et al, 2022; Müller et al, 2022). Significantly increased soil NH 4 + –N was shown in the DA and PA treatments (Table 1), and these results might be accounted for different reasons: (1) nitrification inhibitor DMPP inhibited soil nitrification and then directly enhanced soil NH 4 + –N but decreased NO 3 − –N content (Hargreaves et al, 2021; Jin et al, 2022; Meng et al, 2021; Wang, Liu, et al, 2021), and previous studies suggested that the PSB inoculation also contributed to organic N mineralization (Dong et al, 2022; Jatana et al, 2020; Liu, Bai, Wang, Zhang, et al, 2022); (2) organic residues decomposition by soil microbes might elevate soil NH 4 + –N content (Zhang et al, 2021), which was supported by positive correlations between the soil NH 4 + –N and Peptostreptococcus , Entransia , and Candidatus_Nucleicultrix in this study (Figure 6); and (3) enhanced soil protease activity played a functional role for improving soil NH 4 + –N content (Greenfield et al, 2021).…”

“…Mineral nitrogen (N) is closely related to microbial activity and agricultural crop yield, but low N nutrition supply generally occurs to impair soil microbial biomass and activity and crop yield (Fan et al, 2022; Müller et al, 2022). Protein is regarded as one of the most abundant organic molecules containing numerous N in the soil, which could be hydrolyzed by protease activity to accelerate N metabolism and cycling (Geisseler et al, 2010; Greenfield et al, 2021).…”