A number of proteins functioning in aerobic respiration are shared between eubacteria and archaea (pictured in the following images) and thus may have existed in the last common ancestor. 


These include cytochrome oxidase (subunits I and II), cytochrome b, Rieske iron-sulphur, blue copper protein, 2Fe-2S and 4Fe-4S ferredoxins, and succinate dehydrogenase (the iron-sulphur subunit) (Castresana, 1995).  While most archea live in anaerobic environments, some are aerobic such as Halobacterium, Thermoplasma, Sulfolobus, and Pyrobaculum. NO reductase is homologous to cytochrome oxidases and denitrification may be the origin of aerobic respiration. (Castresana, 1995).  The Krebs cycle can not only be used for energy, it can generate molecules such as NAD+, NADP+, oxaloacetate, succinyl-CoA, ad α-ketoglutarate.   Some of the enzymes used in the Krebs cycle are known in anaerobic bacteria, such as aconitase and isocitrate dehydrogenase (Baughin, 2002).

     In the mitochondria, pyruvate (the end product of glycolysis) is converted to acetyl CoA.  Acetyl CoA is joined to a starter molecule, oxaloacetate, and over the course of a number of chemical reactions, oxaloacetate is produced once again so that the cycle of reactions can begin again.  The carbon atoms which entered the cycle as part of organic molecules are released as carbon dioxide and hydrogen atoms from these biomolecules (and reacting water molecules) are carried by the coenzymes NAD and FAD to the electron transport system.


1)       Pyruvate Dehydrogenase

     Pyruvate dehydrogenase is one of the most complex enzymes known.  It is 30 nm in diameter (larger than a ribosome) and is composed of 60 subunits.  This enzyme catalyzes the conversion of pyruvate and coenzyme A to carbon dioxide and acetyl coA.  Acetyl coA then enters the citric acid cycle.


2)       Citrate Synthase joins Acetyl CoA and Oxaloacetate to produce Citrate


3)       Aconitase converts Citrate to cis-Aconitate


4)       Aconitase converts cis-Aconitate is converted to Isocitrate


5)       Isocitrate dehydrogenase convertes Isocitrate to a Ketoglutarate


6)       The a-Ketoglutarate dehydrogenase complex converts a-Ketoglutarate to Succinyl CoA


7)       Succinyl CoA Synthase converts Succinyl CoA to Succinate


8)       Succinate Dehydrogenase converts Succinate to Fumarate


9)       Fumarase converts Fumarate to Malate


10)  Malate dehydrogenase converts Malate to Oxaloacetate


     Metal ions and sulfur are important in modern organisms in their redox reactions and seem to play an even greater role in simpler organisms.  Hydrogenases may require nickel, iron, and sulfur; reactions involving carbon monoxide may require nickel; the enzymes of the Krebs cycle may require iron and sulfur; some nitrogenases require iron, sulfur, vandium,and molybdenum; ribonucleotide reductase requires iron, sulfur, and cobalt. (Baltscheffsky, p. 116-8).  Ferrodoxins are small and stable iron and sulfur proteins which transport electrons.  They may have been more important in the redox reactions of primitive cells and were gradually replaced over time.  Aconitase is an iron-sulfur protein while fumarase is an iron-sulfur protein in anaerobic bacteria but not so in aerobic organisms  (Baltscheffsky, p. 46).  Once these ions were involved in metabolism, selective pressure would have existed to maximize efficiency—any organisms which possessed mechanisms to regulate the concentrations of these substances would have an advantage.  To maintain the proper charge on these ions, pH would have to be regulated and this would then involve the regulation of sodium, potassium, and chlorine ion concentrations (Baltscheffsky, p. 116-8).

     Oxygen changed everything.  Increases in oxygen concentration led to decreases in concentrations of methane and hydrogen sulfide.  Metals were present in different charged states (Fe3+ as opposed to Fe2+ for example) than existed prior to the increase in oxygen concentrations.  Calcium and zinc became important and eukaryotes would employ both—calcium for signaling systems and zinc for gene regulation.  Eukaryotes utilize zinc for processes for which prokaryotes use iron.  Calcium, once a poison for prokaryotes, could be utilized and, as cells regulated its concentration, it could even be stored in extracellular materials.  Calcium and calmodulin signalling (in addition to inositol phosphate signaling) occur only in eukaryotes.

    Cholesterol synthesis became significant and eukaryotes (such as Euglena pictured below) incorporated significant amounts of cholesterol into their cell membranes.  This made them stronger but also more flexible, allowing a greater variety of sizes and shapes in addition to endocytosis and phagocytosis (including the capture of future endosymbionts) (Baltscheffsky, p. 123-33).  Steroid metabolism produced vitamin D and a variety of steroid hormones.  Oxygen also increased the presence of halogens; these might have been excreted from the cell as poisons at first, only to be later utilized as hormones as the iodine incorporated into the hormone thyroxine  (Baltscheffsky, p. 134).



     Modern bacteria, such as Rhodospirillum and Paracoccus possess the respiratory chain found in mitochondria.  Instead of hydrogen ions being pumped outside of an inner membrane, they are pumped outside the cell.  (Baltscheffsky, p. 26; Darnell, p. 595).

     In the Krebs Cycle, hydrogen atoms were removed from carbon molecules to produce carbon dioxide which is released as a waste.  These hydrogen atoms (hydrogen ions plus their electrons) are transported to the electron transport system in the inner mitochondrial membrane by the coenzymes NAD and FAD.  As electrons are passed from one element of the electron chain to the next, three components of the electron transport system pump hydrogen ions outside the inner membrane into the intermembrane space (NADH dehydrogenase, the cytochrome b-c1 complex, and the cytochrome c oxidase complex).  Once the electrons are deposited into the matrix, a chemiosmotic gradient is created with hydrogen ions on one side of the inner membrane and electrons on the other.  The hydrogen ions are permitted to flow to the electrons at the ATP synthase where ADP is converted to ATP in the process.  The components of the electron transport system consist of:

 --Cytochrome proteins:  These pigmented proteins contain a heme group and include cytochrome a, a3, b, c, and c1.

--FMN: The flavin mononucleotide (like FAD, the flavin dinucleotide) is derived from vitamin B2 (riboflavin).

--Iron-sulfur proteins

--Ubiquinones: Ubiquinones (also known as coenzyme Q or Q) are non-protein molecules.

     Two of the proteins in the chain contain copper atoms.

The mitochondria of higher eukaryotes possess one cytochrome c while many bacteria have multiple electron carriers which function in different physiological conditions (Myllykallio, 1999).


1) NADH Dehydrogenase

     Three of the four divisions of purple bacteria possess ubiquinone oxidoreductase or Complex 1 (including E. coli).   Some bacteria possess a NADH dehydrogenase II which doesn’t pump protons and archebacteria  may have F420H2:quinone oxidoreductase which may be related to Complex 1. (Baltscheffsky, p. 205-6).  The large subunit of all known hydrogenases are homologous to part of Complex 1.  The proton pump quinone reductase possesses many of the subunits of complex 1 and may represent the ancestral form of it (as well as other complexes in purple and cyanobacteria; cyanobacteria are depicted below) (Baltscheffsky, p. 214-5). 


One subunit is an acyl carrier protein ACP similar to bacterial ACPs used in the metabolism of fatty acids.  All hydrogenases have a common ancestor. (Baltscheffsky, p. 216)


2)       Succinate-CoQ reductase


3) bc1 complex (CoQH2-cytochrome c reductase)

Cytochromes bc1 (purple bacteria) and b6f (of gram + bacteria and cyanobacteria) have heme groups and are descended from a common ancestor.  While eukaryotic bc1 complexes may involve 9-11 subunits, bacterial versions only require 3 subunits to function: cytochrome b, cytochrome c1, and the Rieske iron sulfur protein  (Darnell, p. 599; Baltscheffsky, p. 223). 


4)       Cytochrome C oxidase  Complex

     Cytochrome c oxidase is known from bacteria (such as Rhodobacter).  FixN is the member of the cytochrome c oxidases most unrelated to each other.  With the exception of FixN and 1 archeal enzyme, all have subunits II and III.  In most eukaryotes, the core enzyme is encoded by Cox1-3 by mitochondria (bacterial enzyme); 8 to 11 additional subunits are encoded by eukaryotic nucleus  (Darnell, p. 599; Baltscheffsky, p. 254-5).  Cytochrome oxidase must have preceded the rise in oxygen concentration and this enzyme is known in all 3 domains of living organisms.   Oxygen which rusted iron in fossil sediments (redbeds) are known from 2.4-2.6 billion years ago and the modern concentration of oxygen was reached around 500 million years ago.  The original enzymes may have functioned in denitirification—denitrifying and oxygen reducing chains are similar.  The first oxidase, FixN, reduced oxygen rather than joining two NO  (Baltscheffsky, p. 284-90).  In eukaryotes, 7-13 protein subunits form the cytochrome c oxidase complex. The number and identity of subunits can vary between species and between different tissues (Lenka, 1998). Unlike other mammals, anthropoid primates have undergone a number of changes in the enzyme cytochrome c oxidase enzyme, including changes of 11 of the 22 amino acids which interact with cytochrome c (Schmidt, 2005).


5)       Cytochrome c

6) ATP Synthase: F0 /F1 Complex

     ATPases in every cell.  There are two  kinds: F and V and they share a common ancestor.  F are found in eubacteria and are part of mitochondria and chloroplasts.  V ATPases are found in archebacteria and in the vacuoles of eukaryotes.  In anaerobic bacteria, F use ATP energy to pump H+.  The ancestral protein may have been a monomer.  After gene duplication, some of the subunits lost their catalytic capabilities  (Baltscheffsky, p. 291).   Bacterial F0 particles have 3 subunits (a, b, and c) while eukaryotes have these subunits plus 2-5 more.  Bacterial F1 particles are composed of 5 subunits.


     Eukaryotes (such as the amoeba in the following image) possess electron transport systems in their cell membranes (such as the cytochrome P450 pathway) in addition to those in their mitochondrial membranes (Berry, 2003).


One of the techniques used to determine which genes have had the greatest importance in the evolution of a specific lineage is to examine which genes have undergone an accelerated rate of change compared to homologs in related organisms.  The proteins composing the electron transport chain in advanced primates have experienced this type of positive selection, suggesting that these modifications were important in primate adaptations, such as a large brain with increased oxygen requirements.  Proteins of the electron transport chain have experienced positive selection in the lineage leading to higher apes (such as COX4-1, COX7AH, COX8L, and ISP), apes (COX4-1, COX8L, ISP), catarrhine primates (COX2, COX6B, COX6C, COX7C, CYCS, ISP; COX8H became a pseudogene), anthropoid primates (COX1, COX6B, COX6C, COX7C, COX8L, CYCS, CYB,ISP) and primates (COX8H) (Grossman, 2004).