1. Nitrogen Fixation
Our goal is to learn how N2 an inert gas, becomes part of the structure of organic molecules
Secondly, to study the function of nitrogen
compounds in plants and bacteria
To study the nitrogenase complex and
learn its secrets for fixing nitrogen

2. Haber Process How Powerful is Nitrogen Gas? 300 atm N2(g) + 3H2(g)
2NH3(g) + 92 kJ
450oC
(Super heated Steam)

3. Why is Nitrogen so Hard to Split?
Triple bond
945 kJ per mole
(C-C Bond is 347 kJ per mole) N
+ 2e + 2H+
HN=NH
HN=NH
+ 2e + 2H+
H2N-NH2
H2N-NH2
+ 2e + 2H+
2NH3

4. Valence State is a Key to the Chemistry of N+5
HNO3
Nitrates +4
NO2(g)
OXIDATION
Nitrites
HNO2 +3 +2 NO
Nitric oxide +1
N2O(g)
Diatomic N2 -1
HN=NH
REDUCTION -2
H2N-NH2 -3
NH3
Ammonia

5. Nitrogen Compounds in Plants and Bacteria
Nitrite Reduction
Return
nitrogen compounds to N2
Nitrate
Reduction
Denitrification
NO3
NO2
N2(g)
NH3
Energy
Energy
Nitrification
(Soil bacteria)
N2 fixation (reduction)
Plants produce NH3 by reducing nitrates and nitrites
Soil bacteria derive energy by oxidizing NH3

6. Nitrogen Metabolism by Species
Bacteria Plants Animals
Nitrogen fixation
Yes No No
Nitrate Reduction
Yes
Yes No
Nitrite Reduction
Yes
Yes No
Nitrification
Yes No No
Denitrification
Yes No No
NH3 Assimilation
Yes
Yes
Yes
Amino Acid Synthesis
Yes
Yes
Yes

7. Remarkable Nitrogenase
Biological Reaction
N2 + 8H+ + 8e- + 16ATP + 16H2O
2NH3 + 16ADP + 16Pi + H2 + 16H+
Nitrogenase Complex (nif gene cluster)
Nitrogenase (Mo, Fe protein)
Nitrogenase Reductase (Fe protein)

8. Nitrogenase Dialogue Where do we find Nitrogenase?
Nitrogenase is present in certain bacterial species referred
to a diazotrophs. The legumes (peas, beans, alfalfa, etc.)
have root nodules infested with the nitrogen fixing
bacteria, Rhizobium. Free living bacteria such as
azobacter vinelandii can also fix nitrogen.
What is meant by symbiosis in nitrogen fixation?
Symbiosis means living together for mutual benefit. The
bacteria provide the plant with NH3 and the plant prevents
oxygen from interfering with the fixation reaction.

9. What interferes with nitrogen fixation?
Oxygen is the major factor. Nitrogen fixation can take
place only in the total absence of O2
How does a plant overcome oxygen interference?
By synthesizing a heme protein, leghemoglobin, which
like other hemoglobins, has a high affinity for binding
oxygen….what is called “sequestering” the oxygen.
Why such as funny name?
The name gives the origin of the protein. It is made in
legumes, hence leghemoglobin. The heme group of the
hemoglobin is made in the bacteria.

10. SHOW ME Why are there two components in the nitrogenase complex
One component holds the N2 molecules, the
other passes electrons. Nitrogenase is a two-step energy
machine in which electrons are boosted to a higher
reducing potential (Nitrogenase reductase or Fe-protein)
and then passed to the N2 bound to the MoFe-protein
SHOW ME

11. Nitrogenase MoFe-Protein Reductase Fe-Protein N = DinitrogenaseH N = 2 F d e R A T P D . 9 4 3 c t i v a
Electron Transfer “Pump”
Dinitrogenase
Dinitrogenase Reductase
e + H+
Nitrogenase
MoFe-Protein
Reductase
Fe-Protein

12. Of the 3, the most versatile is glutamine
The only inorganic
nitrogen source for
mammals is NH3
NH3
3 Gateways to
Biological Molecules
CO2
ATP
a-Ketoglutarate
Aspartate
Glutamate
Carbamoyl
phosphate
Asparagine
Glutamate
Glutamine
None
Purine nucleotides,
Cytidine nucleotides
Amino sugars,
Tryptophan, Histidine
Pyrimidine nucleotides
Arginine
Urea
Other
amino
acids
Of the 3, the most versatile is glutamine

13. Rapid Quiz
What key biocompound allows a bacterium, a fungus, or a hippopotamus to convert carbohydrates and amino acids into fat?
Based on your answer, show how the carbons in the amino acid alanine and glucose become carbons in cholesterol
Outline a path to show how the carbons in phenylalanine become the carbons in stearic acid and glucose.

14. Answer Aminoacids Carbohydrates Acetyl-CoA Fatty acids Cholesterol
Acylglycerolphosphates
Triacylglycerols
Sphingolipids

15. Acetyl-CoA Alanine Glucose Stearate Pyruvate HMG-CoA Mevalonate
Isopentenyl-PPi
Cholesterol

16. Phenylalanine
Fumarate
Acetoacetate
Tyrosine
L-malate
Acetyl-CoA
p-hydroxyphenylpyruvate
Malonyl-CoA
OAA
Homogentisate
PEP
Palmitate
Maleyl-acetoacetate
Gluconeogenesis
Stearate
Fumaryl-acetoacetate
Glucose