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VUMIE Bacterial Physiology:  Oxidative Phosphorylation

Think back to your chemistry courses and the definitions for oxidation and phosphorylation.  One definition for oxidation is loss of electrons.  In oxidation-reduction (O-R or redox) reactions, one substance loses the electrons (and is oxidized) and another gains the electrons (and is reduced).

Glycolysis and the TCA Cycle are responsible for completely oxidizing nutrients like sugars and amino acids into waste material called carbon dioxide (CO2).  Oxidation occurs when electrons are removed in pairs.  Because two protons (H+) are also removed in the process, oxidation is sometimes called dehydrogenation (hydrogen atoms are an electron and a proton).  Enzymes that do this are called oxidoreductases (as in, "oxidation-reduction") or dehydrogenases. A hydrogen is composed of a proton and an electron.

So where do the electrons and protons go?  If something is oxidized, something else must be reduced!  The electrons and protons are carried by vitamin-derived molecules called NAD and FAD (think UPS trucks for the cell) to the cytochromes for disposal.  This process is called oxidative phosphorylation (Ox-Phos) because it results in the addition of a phosphate to ADP to make ATP.

Remember the role of cytochromes in making ATP.  They accept electrons and pass them along from cytochrome to cytochrome in the Electron Transport System.  While cytochromes can pass along electrons, they cannot deal with the protons.  And so some of the energy of the electrons is lost as the cell deals with the protons by passing them outside of the membrane.  Each pass of an electron represents a bit of dissiplation of energy and more protons ejected through the membrane.  The process of electron transport is completed when the electrons are finally disposed of in the "final electron acceptor", an inorganic molecule that receives the electrons to create a byproduct of growth.  The enzyme that does this is the terminal cytochrome, sometimes called cytochrome oxidase.  In humans and many microbes, that final electron acceptor into which oxidase places the electrons is oxygen.  We need oxygen for life because with out it, there is no place to dispose of the electrons from metabolism and this stops metabolism.  No oxygen, no metabolism.

Bottom line, electron transport disposes of the electrons from metabolism and builds a proton gradient across a membrane (more protons on one side than the other).

Along with electron transport, there is a second process occurring in Ox-Phos.  You may have noticed in the discussion of electron transport that no ATP was made.  That is because it is the product of the second process of Ox-Phos, called chemiosmosis.  When the proton gradient is formed, there is pressure for the protons to cross back through the membrane to equalize concentrations on both sides (think about the sodium-potassium pump and membrane polarization in nerve cells).  When there are higher concentrations on one side of a membrane, ions will try to cross to equalize things.  This is accomplished for protons via the ATPase of the cell.  Protons are allowed to pass through, and as long as they do, the ATPase adds phosphate to ADP to make ATP.  So chemiosmosis takes the proton gradient generated by electron transport and uses it to make ATP.

So how does this translate into something we can study in a micro lab?  There are biochemical tests used in identifying bacteria that look at some of the aspects of energy production metabolism.  In VUMIEtm 2012 and MDM Exercise 9 "Oxidase and O-F Glucose", you will look at the tests used to determine whether the unknown being studied possesses the terminal oxidase cytochrome.  You will also look at whether the microbe is capable of Ox-Phos (in which case it would be called an oxidizer) or instead is forced to take an alternate path called fermentation (in which case it would be called a fermenter).  Fermentation does not result in complete oxidation of the nutrients, but instead is a partial oxidation that does not rely on the TCA Cycle or Ox-Phos.

In addition to Exercise 9, there is another biochemical test that can have relevance to Ox-Phos in a bacterial cell.  In electron transport for aerobic respiration, molecular oxygen (O2) is the final electron acceptor. Electrons and protons are added to make water (H2O).  From your chemistry, you probably noticed that in balancing the equation you end up with an atom of oxygen (O-) left over.  This is called singlet oxygen and it is very dangerous to a cell.  Sometimes it combines with another molecule of oxygen or water to make ozone or hydrogen peroxide, both of which are toxic to cells.  Some bacteria possess protective enzymes that detoxify these molecules.  One such enzyme is catalase, which converts hydrogen peroxide (2 H2O2) into water (H2O) and molecular oxygen (O2)  Exercise 16 "Catalase Test" explains all this and provides an opportunity to see how it is done and why it is useful.

When you have completed these two exercises, you may be asked to take a quiz over this material.