Biochemistry Online: An Approach Based on Chemical Logic

Biochemistry Online





Learning Goals/Objectives for Chapter 8A:  After class and this reading, students will be able to

  • explain why oxidation reactions with ground state dioxygen have a high enough activation energy to make the reactions, although thermodynamically favored, kinetically slow
  • explain, using molecular orbital diagrams the difference between triplet and singlet dioxygen
  • using molecular orbital diagrams and Lewis structures, describe the chemical properties of the reduction products of dioxygen (superoxide, peroxide, and water)
  • explain the ways that biological systems use to enhance dioxygen activity and reduce the effects of reactive oxygen species (ROS) such as superoxide and peroxide
  •  write chemical reactions and mechanisms when appropriate for some reactions of triplet and singlet dioxygen, superoxide, peroxide and the hydroxy free radical
  • describe typical reaction of ROS with lipids, proteins, and nucleic acids and data to support the involvement of ROS in complex diseases and aging.
  • Briefly contrast the production and biological activities of ROS and reactive nitrogen intermediates (RNIs)

A8.  Just Say NO - The Chemistry of Nitric Oxide

In the last decade, the role of another gaseous free radical, nitric oxide (.NO) has become apparent.  This molecule is synthesized biologically through the action of an inducible heme enzyme, nitric oxide synthase, which forms NO by the oxidation by dioxygen of the guanidino group of Arg, which gets converted to citrulline.  .NO is soluble and can diffuse through cell membranes into the cytoplasm, where it has a myriad of effects in signal transduction pathways.  However, it can also be metabolized to form reactive nitrogen intermediates (much as with dioxygen) which can be deleterious to the body, if they damage native biomolecules, or advantageous, when they are used by immune cells like macrophages in the destruction of engulfed bacteria.

A molecular orbital diagram of .NO shows it to have a bond order of 2.5 and one unpaired electron in a π2p* antibonding orbital (hence the notation .NO).

Figure:  Molecular Orbital Diagram of NO

  Here are some of the relevant reactions of NO and its reactive nitrogen intermediates (RNI):

Macrophages make use of RNIs in the killing of engulfed bacteria.  How does one of humankinds greatest foes, Mycobacteria tuberculosis, the causative agent of tuberculosis, avoid this killing mechanism?  It is estimated that the bacteria resides persistently in latent form in 2 billion people.  If it becomes activated, it becomes one of the greatest killers.  Consider the following table:

Killer Diseases through time (Scientist, June 2, 2003)

Historic Pandemics cases deaths
Justinian Plaque (6th centr.) 142 million (on 70% mort) about 100 mill
China Plaque (Bubonic) 3rd Pandemic (1896-1930) 30 milion 12 milion
Spanish flu 1918-19 1 billion 21 million

Pandemics Today

Pandemics Today cases/yr deaths/yr
Malaria 300-500 mil 1 million
TB 8 mill 2 mill
AIDS 6 million 3 mill

People infected with the bacteria but without clinical symptoms must mount a sufficient enough immune response to restrain proliferation of the bacteria, but not enough to clear it from the body.  Immune-compromised people (transplant recipients  taking anti-rejection drugs or AIDS patients) have more difficulty in holding the bacteria at bay. One immune response mediator which restrains  bacterial growth is the soluble protein interferon γ (a protein cytokine released by immune cells).  This protein induces synthesis of nitric oxide synthase, producing .NO, which through the reactions listed above, can damage bacterial macromolecules.

Bacteria are "digested" in acidic phagosomes of the macrophage.  Nitrite formed in the acid conditions from .NO (generated by inducible NO synthase) forms .NO2 and peroxynitrite which have antibacterial properties (better than anti-Tb drugs).  Mutant mice that can not synthesize inducible NO synthase have little defense against the bacteria.  Darwin e al. exposed the bacteria to sources of nitrite at pH conditions typical of macrophage phagosomes and found mutants sensitive to the RNIs.  The mutations involved protein of the bacterial proteasome, a complex multi-protein complex which proteolyzes unwanted (presumably damaged) cellular proteins. 

Bacterial genes encoding proteins associated with the bacterial proteasome seem to confer resistance to the effects of macrophage-inducted RNI production.  The macrophage proteasome, like other eukaryotic proteasomes, is a cytoplasmic protein complex which degrades damaged cytoplasmic proteins.  Although the mechanism is uncertain, the bacterial proteasome may rid the bacteria of nitrated or otherwise oxidized (damaged) proteins or remove the nitrate and facilitate refolding of the damaged protein. 

Peroxynitrite in health and disease

Nitric Oxide Synthases: Structure, Function, and Inhibition


Return to Chapter 8A:  The Chemistry of Dioxygen Sections

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Archived version of full Chapter 8A:  The Chemistry of Dioxygen


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