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    21 - N Fixation PDF

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    shubham
    Biological nitrogen fixation is the process by which nitrogen-fixing bacteria and archaea convert atmospheric nitrogen into ammonia. This process is essential as it is responsible for about 70% of the nitrogen entering the biosphere. The ability to fix nitrogen is restricted to certain prokaryotes, with the main nitrogen-fixing bacteria being rhizobia, azotobacteria, and cyanobacteria. Rhizobia have the ability to form symbiotic relationships with legumes, living in root nodules and fixing nitrogen for the plant in exchange for carbon sources. The nitrogenase enzyme is key in this nitrogen fixation process, reducing nitrogen gas into ammonia through an energy-intensive process requiring ATP.

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    21 - N Fixation PDF

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    shubham
    135 views7 pages
    Biological nitrogen fixation is the process by which nitrogen-fixing bacteria and archaea convert atmospheric nitrogen into ammonia. This process is essential as it is responsible for about 70% of the nitrogen entering the biosphere. The ability to fix nitrogen is restricted to certain prokaryotes, with the main nitrogen-fixing bacteria being rhizobia, azotobacteria, and cyanobacteria. Rhizobia have the ability to form symbiotic relationships with legumes, living in root nodules and fixing nitrogen for the plant in exchange for carbon sources. The nitrogenase enzyme is key in this nitrogen fixation process, reducing nitrogen gas into ammonia through an energy-intensive process requiring ATP.

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    Biological nitrogen fixation is the process by which nitrogen-fixing bacteria and archaea convert atmospheric nitrogen into ammonia. This process is essential as it is responsible for about 70% of the nitrogen entering the biosphere. The ability to fix nitrogen is restricted to certain prokaryotes, with the main nitrogen-fixing bacteria being rhizobia, azotobacteria, and cyanobacteria. Rhizobia have the ability to form symbiotic relationships with legumes, living in root nodules and fixing nitrogen for the plant in exchange for carbon sources. The nitrogenase enzyme is key in this nitrogen fixation process, reducing nitrogen gas into ammonia through an energy-intensive process requiring ATP.

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    135 views7 pages

    21 - N Fixation PDF

    Uploaded by

    shubham
    Biological nitrogen fixation is the process by which nitrogen-fixing bacteria and archaea convert atmospheric nitrogen into ammonia. This process is essential as it is responsible for about 70% of the nitrogen entering the biosphere. The ability to fix nitrogen is restricted to certain prokaryotes, with the main nitrogen-fixing bacteria being rhizobia, azotobacteria, and cyanobacteria. Rhizobia have the ability to form symbiotic relationships with legumes, living in root nodules and fixing nitrogen for the plant in exchange for carbon sources. The nitrogenase enzyme is key in this nitrogen fixation process, reducing nitrogen gas into ammonia through an energy-intensive process requiring ATP.

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    Biological N2 fixation (BNF)

    Fixation of elemental nitrogen in the atmosphere by the microorganisms through a


    reductive process into ammonia is called as BNF. A variety of prokaryotic organisms have the
    ability to reduce the atmosphere N2. BNF accounts for about 70% of the total N fixed in the
    biosphere. The ability to reduce atmosphere N is restricted only to bacteria, belonging to
    diverse groups. The root nodule associations were the first to be recognized for their ability to
    fix atmosphere N2. Rhizobia are the first group of organism realized for its potential of nitrogen
    fixation.
    Nitrogen fixing bacteria
    Nitrogen fixing bacteria are classified according to their mode of fixation.
    1. Free living N fixers: fix mol. N2 to cellular N independently of other organisms.
    2. Associative N fixers
    3. Endophytic N fixers
    4. Symbiotic N fixers
    Rhizobium is predominant symbiotic N2 fixing bacterium. Boussingault showed that
    leguminous plant can fix atmosphere N2. Then Hellriegel and Wilfarth – proved that N2 is fixed
    by certain bacteria living in root nodules of leguminous plants. Latter isolated in pure culture by
    Beijerinck. Winogradsky isolated Clostridium pasteurianum, which is an anaerobic N2 fixer.
    Beijerinck isolated Azotobacter as a free living aerobic N2 fixing organism.
    Cross inoculation groups of rhizobium (CIG)
    It (CIG) refers the groups of leguminous plants that will develop effective nodules when
    inoculated with the rhizobia obtained from the nodules from any member of that legume group.
    I. Rhizobium
    1. Rhizobium leguminosarum CIG Host it can nodulate
    bv. Viceae Pea Peas,lenfils, vicia
    bv. Phaseoli Bean Phaseolus spp
    bv. Trifoli Clover Trifolium spp
    2. R. meliloti Alfalfa Alfalfa, clover, fenugreek
    3. R. loti Lotus Trifoli, lupine,
    4. R. fredii Soybean Soybean
    5. R. spp Cowpea group Vigna, Arachis, Cajanus, Dolichus,
    Sesbania, Acacia, Prosopis, green
    gram and blackgram
    6. R. sp Chickpea group Chickpea
    II. Bradyrhizobium
    B. japonicum Soybean Soybean
    B. spp Cowpea group Cowpea group plants

    III. Azorhizobium - Stem nodulating – one. Nodulates Sesbania rostrata.


    IV. Photorhizobia - Nodulants aeschynomene sp.
    V. Sinorhizobium - fast growing soybean nodulator

    I. Biological nitrogen fixation


    Free living nitrogen fixers
    Azotobacter (Aerobic) - A. chroococcum, A. vinelandii, A. beijerinkii, A. paspali, A. agilis,
    A. insignis, A. macrocytogens
    Beijerinckia
    Clostridium – Anaerobic
    Cyanobacteria (BGA) - Anabaena, Nostoc, Cylindrospermum, Rivularia, Oscillatoria,
    Plectonema, Aphanothece, Lyngbya, Scytonema, Calothrix etc.
    II. Associative symbiotic nitrogen fixer
    Azospirillum - A. Lipoferu, A. Brasilense, A. Amazonense, A. Halopareferans, A. irkense,
    A.largomobilis
    Herbaspirillum
    III. Endophytic nitrogen fixer
    Gluconacetobacter diazotrophicus
    IV. Symbiotic nitrogen fixers
    Rhizobium (Rhizobium – legume association)
    Bradyrhizobium (Bradyrhizobium – soybean association)
    Azorhizobium (Azorhizobium- Sesbania rostrata association)
    Anabaena azollae (Azolla – Anabaena association)
    Frankia (Frankia – Casuarina association)
    Species of Azolla: A. pinnata, A. filiculoides, A. Microphylla, A. Caroliniana,
    A. Mexicana, A. nilotica
    Nitrogen fixation:
    Process of N2 fixation
    The process of N2 fixation is mediated by the enzyme called nitrogenase (which
    mediates the reduction of N2 to ammonia) first, this enzyme was extracted from the anaerobic
    di nitrogen fixer Clostridum pasteurianum. Latter, this has been isolated form most other N2
    fixing bacteria.
    The mechanism of N2 fixation appears to be quite similar in most N2 fixing prokaryotes.
    The enzyme has been fairly well characterized and the enzymes from these different systems
    share common properties allowing a unified single description of nitrogenase.
    Nitrogenase
    Nitrogeanse is a functional enzyme which reduces N2 to ammonia and depends on
    energy source from ATP. The nitrogenase has two components one containing Mo-Fe,
    designated as Mo – Fe protein and the other Fe protein. Two components are necessary for
    the nitrogenase activity.
    Mo-Fe protein
    Consists of 4 subunits and having the molecular not of 22,0000 or 270,000 daltons and it
    is the big component.
    Fe-protein
    Smaller component, contains 2 subunits, molecular weight 60,000 daltons.
    Ammonia is the end product of N2 fixation. The over all reaction is as follows.
    ATP
    N2 + 3H2 2 NH3
    General pathway of N2 fixation
    This process requires a source of ATP and reductants, which are provided by
    photosynthesis. 16 molecules of ATP are required to fix a molecule of N2.

    ATP ATP ADP


    generation
    N=N
    Energy source Electron Nitrogenase
    carriers complex
    NH3

    Nitrogenase can also reduce C2 H2 → C2H4


    Hydrolysis of ATP into ADP with electron transfer from a reduced electron donor
    (Ferridoxin, Flavodoxin) is coupled to reduce N2 to 2NH3. The ammonia is the first stable
    product of fixation and it is assimilated by GS-GOGAT pathway.

    NADH
    NADPH e- e- e- e-
    Ferridoxin Flarodoxin Ubiquinone
    APP
    EMP e- Carrier proteins NII NI
    EDP
    Glycolysis

    Nitrogenase is O2 labile various protection mechanism are operating in different N2


    fixing systems.

    N2 Fixation
    Reductants
    NADH
    NADPH
    e- - -
    Ferridoxin e Flavodoxin e Ubiquinone
    ATP e-
    ATP
    Glycolysis Nitrogenase H+
    EMP C2H2
    EDP
    N2

    II NH3
    I
    C2H4
    ADP +Pi

    H2

    Mechanism
    Reduction takes place on the surface of the enzyme
    • Six electrons are required to reduce one mole of N to two moles of ammonia.
    N2 + 8H+ +8e- +16 ATP -------2 NH3+16 ADP+ 2H+ +16pi
    It is postulated that, atoms of N2 are
    separated thr’h charge in the valency Reduction process
    of metal ion (mo) bound to the N2 Dinitrogen
    enzyme involved in reduction of N2. 2H+, 2e-
    For every electron transfer, 2 ATP HN = HN Diimide
    mole are required.
    2H+, 2e-
    Hydrogenase -Uptake hydrogenase
    H2N- NH2 Hydrazine
    (HUP+) converts the release d
    hydrogen during N2 fixation ,and 2H+, 2e-
    cycled back the Hydrogen for energy HO2HN- NH2OH Hydroxyl amide
    generation.by this they contribute 9-
    2H+, 2e-
    10 %ATP requirement for N2 fixation
    process. 2 NH3 Ammonia
    Factors affecting N2 fixation
    1. Presence of nitrate or ammonium : More N2, No, N2 fixation
    2. Presence of certain inorganic substances
    Ca, Co, Mo – influence N2 fixation along with P
    3. Availability of energy source – addn. of C source increase N2 fixation
    4. pH : Neutral – favours Azotobacter – Acidic- Beijerinkia
    5. Soil moisture : Adequate is good for fixation
    6. Temperature : Mesophilic – 30°C.
    The energy requirement for BNF is very high and it is a major factor determines the
    amount of N2 fixed. In, Azotobacter the rate depends on amount of available carbon. In
    symbiotic N2 fixers since photosynthesis is the ultimate source of energy the rate of N2 fixation
    is influenced by the factors that effect photosynthesis and rate of translocating photosynthates
    to the N2 fixing system.
    Nitrogenase protection mechansims
    1. Leghaemoglobin scavenges O2 to protect nitrogenase in legume rhizobium symbiosis
    2. Confirmatory protection in Azotobacter as well as the higher respiratory rate.
    3. Thick walls of Heterocyst protect O2 in BGA, since Nitrogenase are present in the
    heterocyst.
    4. Microaerophilic nature in Azospirillum
    Losses of N by non biological ways
    Leaching
    20 to 50%of fertilizer N. The most striking loss of N in rice soils where more than half of
    the fertilizer N applied get lost through leaching.
    Volatilization
    Another factor is the volatalizaiotn of ammonia in soil 5-20%.
    Fixation of ammonium in soils is the minor contributory factor to overall loss of N2 available for
    plant growth.
    Such losses of N by physical causes and by nitrification and denitirfication process can
    be controlled by the application of certain chemicals. Some chemicals have been designed to
    control the rate of release of nutrient from nitrogenous fertilizers, while others retard
    nitrification in soil by controlling the activity of nitrifying bacteria.
    a. Controlled release fertilizers
    Urea from
    Fertilizers, sparingly soluble in water can regulate
    isobutyeldene diurea the release of N from fertilizers
    Crotonilidene diurea
    S coated urea
    b. Nitrification inhibitors
    These are substituted with pyridines, pyrimidines, anilines and isothiocyanates,
    Examples: 2 chloro 6 (tricholormethyl) – pyridine – (N serve ), 2 amino 4 chloro 6 methyl
    pyridine –( AM.)
    N serve inhibits the growth of Nitrosomonas europea and N. agilis.
    The seeds of neem conain lipid associates act as nitrification inhibitors and there by
    increases the efficiency of urea fertilizers.
    Ammonia assimilation
    N2 fixation results in NH4 formation which reacts with organic acids and form amino
    acids which is mediated by ammonia assimilating enzyme.
    GS – Glutamine synthetase, GOGAT – Glutamate synthese
    GDH – Glutamate dehydrogenase
    Genetics
    Nif genes are responsible for N2 fixation.
    Nif genes are 22, which are located in 7 or 8 clusters in a 24 kb fragment.

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