Nitrite oxidation (Nxr)

• Page authors: PRIMO
• Responsible curator: Hagen Buck-Wiese

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Nitrite oxidation rate (Nxr proteomics)
Approach: targeted proteomics of Nxr + Michaelis–Menten kinetics
Context: environmental collection
Spatial scale: point sample
Temporal scale: hours to days
Units: nmol NO3- d-1
Community captured: nitrite-oxidising bacteria (NOB)
Co-measurements: specific activity of purified Nxr; Michaelis–Menten kinetic parameters from whole live cells

Method Overview

Nitrite oxidation rates are estimated from the concentration of nitrite oxidoreductase (Nxr) enzyme in field samples combined with its kinetic parameters. Nxr is quantified by targeted proteomics: large-volume water samples (300–500 L, typically from pumping systems) are filtered to concentrate NOB biomass, proteins are extracted and digested, and Nxr-specific peptides are quantified by selected-reaction monitoring (SRM) mass spectrometry using isotope-labelled peptide standards. This gives the in situ Nxr concentration. Nitrite oxidation rates are then estimated from the Michaelis–Menten equation using the measured Nxr concentration, in situ [NO₂^-], and kinetic parameters (V_max, K_m) derived from pure-culture or whole-cell experiments.

Scale of measurement

Very large sample volumes are required (300–500 L) to concentrate sufficient biomass of NOB for protein detection, limiting spatial resolution. Temporal resolution is set by sample collection logistics rather than the incubation itself.

Data generated

In situ nitrite oxidation rates in nmol NO₃^- d^-1, derived from enzyme concentration and kinetic parameters.

Units & currency

Units are nmol NO₃^- d^-1. The currency is nitrogen (gene/transcript/protein abundance; rate estimated from enzyme concentration).

Sample size

Typical samples are 300–500 L (large-volume pumping).

Limitations

The approach assumes substrate limitation and applies kinetic parameters measured from pure cultures or whole cells to mixed field communities, where the physiological state and kinetic parameters of individual NOB populations may differ substantially. The specific activity of purified Nxr and Michaelis–Menten parameters derived from isolated organisms may not accurately represent the diverse Nxr variants in environmental communities. High sample volume requirements restrict applicability.

Example Applications & Protocols

Recent applications

• Saito et al. (2020) Needles in the blue sea: sub-species specificity in targeted protein biomarker analyses

Common calculations/conversions

• Nitrite oxidation rate = [Nxr] × V_max × [NO₂^-] / (K_m + [NO₂^-]).