Nitrate reductase

Key takeaways

Nitrate reductase belongs to biology; place it in that map before memorising details.
Learn the definition first, then one example that makes the definition concrete.
Connect Nitrate reductase to a quantity you can measure, compute or draw — that is where exam questions come from.
Reproduce the core statement of Nitrate reductase from memory before moving on to harder problems.

Reference excerpt

Nitrate reductases are molybdoenzymes that reduce nitrate (NO−3) to nitrite (NO−2). This reaction is critical for the production of protein in most crop plants, as nitrate is the predominant source of nitrogen in fertilized soils.

Types

Eukaryotic Eukaryotic nitrate reductases are part of the sulfite oxidase family of molybdoenzymes. They transfer electrons from NADH or NADPH to nitrate.

Prokaryotic Prokaryotic nitrate reductases belong to the DMSO reductase family of molybdoenzymes and have been classified into three groups, assimilatory nitrate reductases (Nas), respiratory nitrate reductase (Nar), and periplasmic nitrate reductases (Nap). The active site of these enzymes is a molybdenum ion that is bound to the four thiolate functional groups of two pterin molecules. The coordination sphere of the molybdenum ion is completed by one amino-acid side chain and oxygen and/or sulfur ligands. In Nap, the molybdenum is covalently attached to the protein by a cysteine side chain, and an aspartate side chain in Nar.

Structure Prokaryotic nitrate reductases have two major types, transmembrane nitrate reductases (NAR) and periplasmic nitrate reductases (NAP). NAR allows for proton translocation across the cellular membrane and can contribute to the generation of ATP by the proton motive force. NAP cannot do so. The transmembrane respiratory nitrate reductase is composed of three subunits; an 1 alpha, 1 beta and 2 gamma. It can substitute for the NRA enzyme in Escherichia coli, allowing it to use nitrate as an electron acceptor for anaerobic respiration. A transmembrane nitrate reductase that can function as a proton pump (similar to the case of anaerobic respiration) has been discovered in the diatom Thalassiosira weissflogii. The nitrate reductase of higher plants, algae, and fungi is a homodimeric cytosolic protein with five conserved domains in each monomer: 1) an Mo-MPT domain that contains the single molybdopterin cofactor, 2) a dimer interface domain, 3) a cytochrome b domain, and 4) an NADH-binding domain that combines with 5) an FAD-binding domain to form the cytochrome b reductase fragment. There exists a Glycophosphatidylinositol-anchored variant that is found on the outer face of the plasma membrane. Its function is not clear.

Mechanism In prokaryotic periplasmic nitrate reductase, the nitrate anion binds to Mo(IV). Oxygen transfer yields an Mo(VI) oxo intermediate with release of nitrite. Reduction of the Mo oxide and protonolysis removes the oxo group, regenerating Mo(IV). Similar to the prokaryotic nitrate reduction mechanism, in eukaryotic nitrate reductase, an oxygen in nitrate binds to Mo in the +4 oxidation state, displacing a hydroxide ion. Then the Mo d-orbital electrons flip over, creating a multiple bond between Mo(VI) and that oxygen, ejecting nitrite. The Mo(VI) double bond to oxygen is reduced by NAD(P)H passed through the intramolecular transport chain.

Regulation Nitrate reductase (NR) is regulated at the transcriptional and translational levels induced by light, nitrate, and possibly a negative feedback mechanism. First, nitrate assimilation is initiated by the uptake of nitrate from the root system, reduced to nitrite by nitrate reductase, and then nitrite is reduced to ammonia by nitrite reductase. Ammonia then goes into the GS-GOGAT pathway to be incorporated into amino acids. When the plant is under stress, instead of reducing nitrate via NR to be incorporated into amino acids, the nitrate is reduced to nitric oxide which can have many damaging effects on the plant. Thus, the importance of regulating nitrate reductase activity is to limit the amount of nitric oxide being produced.

Inactivation of nitrate reductase The inactivation of nitrate reductase has many steps and many different signals that aid in the inactivation of the enzyme. Specifically in spinach, the very first step of nitrate reductase inactivation is the phosphorylation of NR on the 543-serine residue. The very last step of nitrate reductase inactivation is the binding of the 14-3-3 adapter protein, which is initiated by the presence of Mg2+ and Ca2+. Higher plants and some algae post-translationally regulate NR by phosphorylation of serine residues and subsequent binding of a 14-3-3 protein.

Anoxic conditions Studies were done measuring the nitrate uptake and nitrate reductase activity in anoxic conditions to see if there was a difference in activity level and tolerance to anoxia. These studies found that nitrate reductase, in anoxic conditions improves the plants tolerance to being less aerated. This increased activity of nitrate reductase was also related to an increase in nitrite release in the roots. The results of this study showed that the dramatic increase in nitrate reductase in anoxic conditions can be directly attributed to the anoxic conditions inducing the dissociation of 14-3-3 protein from NR and the dephosphorylation of the nitrate reductase.

Applications Nitrate reductase activity can be used as a biochemical tool for predicting grain yield and grain protein production. Nitrate reductase can be used to test nitrate concentrations in biofluids. Nitrate reductase promotes amino acid production in tea leaves. Under south Indian conditions, it is reported that tea plants sprayed with various micronutrients (like Zn, Mn and B) along with Mo enhanced the amino acid content of tea shoots and also the crop yield.

References

External links UMich Orientation of Proteins in Membranes protein/pdbid-1q16 Nitrate+reductases at the U.S. National Library of Medicine Medical Subject Headings (MeSH)

Illustrations

Worked examples

Example 1 — a first encounter with Nitrate reductase

Start with the simplest possible case. Write down what Nitrate reductase claims or describes in one sentence, then invent the smallest concrete situation in which that sentence is true. In biology, the smallest case is usually a single object, a single equation or a single measurement. Check that every symbol or term in your sentence has a meaning in that case.

Example 2 — changing one variable

Take the situation from Example 1 and change exactly one quantity: double it, halve it, or set it to zero. Predict what should happen to Nitrate reductase before you calculate. Comparing your prediction with the result is the fastest way to find out whether you understand the idea or only the words.

Example 3 — an exam-style question

Typical questions about Nitrate reductase ask you to (a) state it precisely, (b) apply it to given data, and (c) explain a limitation. Practise writing all three answers in under five minutes; the third part is what separates a full-mark answer from an average one.

Applications of Nitrate reductase

In research: Nitrate reductase appears in biology research whenever the underlying quantities have to be modelled precisely. Papers usually cite it as a starting assumption and then explore where it breaks down.
In technology and industry: Engineering practice reuses Nitrate reductase in design rules, simulations and safety margins. Knowing the idea lets you read a specification sheet and understand why the numbers look the way they do.
In the classroom: Nitrate reductase is common in secondary-school and first-year university syllabi. It links to neighbouring topics EC 1.7.99, Enzymes, Integral membrane proteins, so understanding it makes those chapters shorter.
In everyday life: Look for Nitrate reductase outside the textbook — in sport, cooking, traffic, electronics or the sky above you. An example you found yourself is remembered far longer than one you were given.

How to study Nitrate reductase in 20 minutes

Read the reference excerpt below once, without taking notes.
Close the page and write down what Nitrate reductase means in your own words.
Compare your version with the excerpt and mark what you missed.
Work through the three examples above with pen and paper.
Explain Nitrate reductase out loud to somebody else — or to Teacher Smith in the lgStudy chat.

Frequently asked questions

What is Nitrate reductase in simple terms?

Nitrate reductases are molybdoenzymes that reduce nitrate (NO−3) to nitrite (NO−2). This reaction is critical for the production of protein in most crop plants, as nitrate is the predominant source of nitrogen in fertilized soils.

Why does Nitrate reductase matter?

Because it connects several biology ideas at once: it gives you a definition you can apply, a quantity you can calculate, and a way to check whether a result is plausible.

How should I study Nitrate reductase?

Read the excerpt, restate it from memory, then work through the examples and applications listed on this page. The five-step study plan above takes about twenty minutes.

What does this page cover?

It gives you a compact reference excerpt plus original lgStudy explanations, examples, applications and study material on Nitrate reductase.