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Nitrogen Fixation

Biological Nitrogen Fixation

Biological nitrogen fixation is accomplished through the catalytic action of an enzyme known as nitrogenase. Nitrogenase consists of two distinct proteins which contain molybdenum, iron, and sulfur. Because the nitrogenase proteins are denatured by exposure to oxygen (O2), they can only operate in an anaerobic environment. Nitrogen fixation using nitrogenase requires rather large inputs of energy to drive the process, equivalent to about 150 calories of energy per mole of nitrogen gas that is fixed. Although nitrogen fixation is energetically expensive, it is nevertheless worthwhile for plants that grow in nitrogen-deficient habitats.

The nitrogenase enzyme can cleave other triple bonds in addition to that of dinitrogen gas, for example, that of acetylene. Most assays of nitrogen fixation rates take advantage of this fact, measuring nitrogenase activity through the rate at which ethylene is generated by the reaction of acetylene with this enzyme.

Only certain microorganisms can synthesize the nitrogenase enzyme. Some of these microbes are free-living in soils and water, while others occur in mutualisms with fungi or plants.

A large number of free-living bacteria of anaerobic environments have the ability to fix nitrogen, including the genus Clostridium. Fewer genera of aerobic bacteria have this ability. Free-living nitrogen fixation occurs most vigorously in habitats containing large quantities of carbon-rich organic debris such as rotting logs, heaps of sawdust, and compost piles.

Blue-green bacteria are free-living, photosynthetic microbes that fix nitrogen in aquatic habitats and moist soil, including the genera Anabaena, Nostoc, and Calothrix. Strains of these microbes also live in mutualisms with certain fungi in a symbiosis known as lichens which have an ability to fix nitrogen. Anabaena and Nostoc also occur in a mutualism with the floating aquatic fern Azolla which can be cultivated on the surface of rice paddies to fix nitrogen at a rate as great as 100-150 kg N/ha.yr, saving on the use of synthetic fertilizer.

Nitrogen-fixing bacteria can also occur in mutualisms with certain vascular plants. The best known of these associations are with species of legumes (order Leguminosae). About one-half of the world's 10,000 species of legumes occur in a nitrogen-fixing mutualism with bacteria in the genus Rhizobium. About 50 of these species are utilized in agriculture, for example, the food crops garden pea (Pisum sativum), broad bean (Vicia faba), and soybean (Glycine max), and the soil conditioners red clover (Trifolium pratense) and alfalfa (Medicago sativa). The strain of Rhizobium is specific to each legume species.

The Rhizobium occurs in specialized nodules on the roots of the legumes. These are developed when the soil-dwelling Rhizobium invades a root hair, stimulating the plant to form a nodule. Nodule development is inhibited in acidic soils and if the concentrations of nitrate in soil are large. To protect the nitrogenase enzyme, the interior of the nodules is anaerobic. This is due in part to the presence of an oxygen-binding pigment known as leghaemoglobin which colors the nodule interior a dark red. The leghaemoglobin also serves as an oxygen carrier important for other aspects of bacterial metabolism, in this way performing a similar function as the haemoglobin of animals.

Several hundred species in other plant families also develop root nodules that can fix nitrogen. The best known of these are the woody plants known as alders (Alnus spp.), which develop nodules containing actinomycetes in the genus Frankia. Red alder (Alnus rubra) reaches tree-size, and its stands can fix hundreds of kilograms of nitrogen per year. Other non-leguminous plants that are known to fix nitrogen include the genera Casuarina, Ceanothus, Myrica, Dryas, and Shepherdia, variously associated with fungi, actinomycetes, or bacteria.

Some species of grasses occur in a relatively loose, non-obligate mutualism with nitrogen-fixing microorganisms. These associative symbioses include those of the crabgrass Digitaria with the bacterium Spirillum and that of another weedy grass, Paspalum, with Azotobacter. Although these nitrogen-fixing symbioses do not involve agriculturally important species of plants, they may nevertheless prove to be significant. Research is being conducted into the possibility of transferring the associative ability of nitrogen fixation of these species into grasses that are important in agriculture, such as corn (Zea mays) and wheat (Triticum aestivum). If this feat of bioengineering could be accomplished, it could lead to significant savings of nitrogen fertilizer which must be manufactured using non-renewable resources.

One last example of a nitrogen-fixing mutualism involves termites. These insects have a bacterium, Enterobacter agglomerans, in their gut that fixes nitrogen. This is important to termites because their diet of cellulose is highly deficient in nitrogen.

See also Nitrogen cycle.



Freedman, B. Environmental Ecology. 2nd ed. San Diego: Academic Press, 1995.

Bill Freedman


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—A relationship between two or more organisms that is mutually beneficial.

Nitrogen fixation (dinitrogen fixation)

—The conversion of atmospheric dinitrogen (i.e., N2) to ammonia or an oxide of nitrogen. This process can occur inorganically at high temperature and pressure and biologically through action of the microbial enzyme, nitrogenase.


—An enzyme synthesized by certain microorganisms that is capable of cleaving the triple bond of nitrogen gas generating ammonium, a biologically useful type of fixed nitrogen.

Additional topics

Science EncyclopediaScience & Philosophy: Nicotinamide adenine dinucleotide phosphate (NADP) to Ockham's razorNitrogen Fixation - Inorganic Nitrogen Fixation, Biological Nitrogen Fixation