Biochemistry - The Chemical Reactions of Living Cells, Volume 2 - D. Metzler 1980

How electrons meet oxygen, how ATP is generated in the process, and other related phenomena
Historical Background

Animal Respiration began to seriously attract the attention of chemists in 1777, when Lavoisier established that food substances undergo slow combustion within the Organism, which he believed occurred in the Blood. Spallanzani's observations, carried out between 1803 and 1807, probably showed for the first time that Tissues are the true site of respiratory processes, but unfortunately his observations failed to attract attention. In 1884, MacMunn discovered that Cells contain heme pigments now known as Cytochromes. However, leading biochemists of the time considered MacMunn's observations to be the result of experimental errors, so that the period of serious Study of the chemistry of Biological Oxidation actually begins only with the present century [1].

The Conclusion that substrates are oxidized via dehydrogenation is commonly associated with the name of Wieland. Between 1912 and 1922, he demonstrated that intracellular "respiration" processes can proceed even in the absence of oxygen, provided that various synthetic Dyes, such as methylene blue, are present. Subsequent experiments (Chap. 8, Sec. 3) led to the isolation of soluble pyridine NUCLEOTIDES and Flavoproteins and to The Development of METABOLISM/2.html">THE CONCEPT OF an Electron Transport Chain. Studying processes at the other end of the Respiratory Chain, Warburg noted (1908) that all aerobic cells contain iron. Moreover, iron-containing charcoal prepared by burning blood was found to catalyze the non-enzymatic oxidation of many substances, whereas iron-free charcoal from cane sugar possessed no such properties. It was discovered that tissue respiration is inhibited by concentrations of cyanide just as low as those required to inhibit the non-enzymatic catalytic action of iron salts. Based on these observations, Warburg suggested in 1925 that aerobic cells contain an iron-containing respiratory enzyme (Atmungsferment); it was later named cytochrome oxidase. This enzyme was shown to be inhibited by carbon monoxide.

It was known that heme-carbon monoxide complexes dissociate under The Influence of light; taking this into account, Warburg and Negelein (1928) recorded the photochemical action spectrum (see note to Chap. 13, Sec. B) for the Reversal of the inhibitory effect of carbon monoxide on the respiration of the Yeast Torula utilis. This spectrum was similar to the absorption spectrum of other heme derivatives. Thus arose the hypothesis that O₂, much like CO, binds to the iron of the heme group that is part of the respiratory enzyme.

Meanwhile, between 1919 and 1925, D. Keilin studied the thoracic Muscles of flies and other insects by examining them under a Microscope equipped with a spectroscopic eyepiece. In doing so, he discovered a pigment with four absorption bands and initially concluded that they were due to some modification of Hemoglobin. However, upon finding the same pigment in fresh baker's yeast, he realized that he was dealing with a new substance. The important role of this substance was established in the following manner (quoting Keilin's recollections [2]):

“One day, while examining a freshly prepared yeast suspension obtained from lumps of yeast paste by vigorous shaking in a test tube with a small amount of Water, I failed to detect the characteristic four-banded absorption spectrum. But no sooner had I removed the suspension from the field of view of the microspectroscope than these four bands suddenly reappeared. I repeated this experiment over and over again, always with the same result: the absorption bands disappeared after shaking the suspension in air and reappeared a few seconds after the shaking was stopped.

I must confess that this first Visual Perception of the intracellular respiratory process was one of the most impressive sights I have ever witnessed in my work. I no longer had any doubt that cytochrome, apart from being universally distributed in nature and completely independent of hemoglobin as a substance, represents an intracellular respiratory enzyme far more important than hemoglobin.”

Keilin soon established that the three absorption bands observed at 604, 564, and 550 nm (a, b, and c) are due to three different pigments, whereas the band at 521 nm turned out to be common to all three. Keilin proposed naming these pigments cytochromes a, b, and c. The concept of Electron transport along the respiratory chain [2] emerged almost immediately after The Role of flavin- and pyridine nucleotide-containing Coenzymes at the substrate dehydrogenation level was established. Hydrogen atoms delivered to these carriers could be used to reduce oxidized cytochromes. The latter could then be oxidized by oxygen with the participation of cytochrome oxidase.

In 1929, Fiske and SubbaRow [2a], interested in the presence of purine compounds in Muscle extracts, discovered and characterized ATP. It was soon demonstrated (primarily through the work of Lundsgaard and Lohmann) that ATP Hydrolysis serves as the energy source for Muscle contraction. Around the same time, it became known that Glycolysis is accompanied by ATP synthesis. The possibility of ATP formation during electron transport became evident after Engelhardt established in 1930 that methylene blue stimulates ATP Synthesis in tissues.

Serious investigation of the Electron Transport Chain and Oxidative Phosphorylation began following the demonstration by Kennedy and Lehninger in 1949 that Cell/35.html">Mitochondria are not only the site of ATP synthesis, but also the site where The Tricarboxylic Acid Cycle and Fatty acid oxidation take place. Chance (1959) completed the development of a new, elegant spectrophotometric apparatus. The data obtained allowed him to postulate the following sequence of carriers in the respiratory chain:

Class="center">Substrate → Pyridine nucleotides → Flavoprotein → Cytochrome b →

→ Cytochrome c → Cytochrome a → Cytochrome a₃→ O₂.----- (10-1)

Since then, several new Components of the respiratory chain have been discovered, most notably ubiquinones and Non-heme iron Proteins.

Some details of this scheme are given below, beginning with a consideration of The properties of Hemoproteins and oxygen itself.