David Keilin

The Leeuwenhoek Lecture: The Problem of Anabiosis or Latent Life: History and Current Concept

The eight previous Leeuwenhoek Lectures covered a great variety of problems is bacteriology and virology, and each of the lecturers paid an enthusiastic tribute Antony van Leeuwenhoek as the founder of microbiology. When the Council honoured me by their invitation to deliver the ninth Leeuwen hoek Lecture I thought that it would be appropriate to devote it to the problem of anabiosis or latent life.

Succinic Dehydrogenase-Cytochrome System of Cells Intracellular Respiratory System Catalysing Aerobic Oxidation of Succinic Acid

Succinic dehydrogenase-cytochrome system, even in the cell-free colloidal preparation, behaves like a true respiratory system of the cell, showing the high catalytic activity and being affected by all the inhibitors in the same way and to the same degree as the respiration of intact cells. The efficiency of this system depends not only on the integrity of all its components but also on that of the colloidal structure which supports them and assures their mutual accessibility. The paper deals with the properties of each of the components of the system and with the mechanism of their interreactions.

Effect of inhibitors on the activity of soluble succinic dehydrogenase and on the reconstitution of the succinic dehydrogenase-cytochrome system from its components

The succinic dehydrogenase-cytochrome system, which catalyzes the oxidation of suecinate to fumarate by molecular oxygen, was successfully reconstituted from soluble succinic dehydrogenase, soluble cytochrome c and a particulate heart-muscle preparation which is deprived of these components but contains cytochromes a3, a, c1 and b and has a strong cytochrome oxidase activity. Although succinic dehydrogenase isolated in a soluble form by Singer and by Wang with their co-workers was identified by them as a flavoprotein compound containing one modified flavin adenine dinucleotide and 2 or 4 iron atoms, neither the function of the iron nor the structure and function of the flavin group have so far been satisfactorily elucidated. Soluble succinic dehydrogenase, unlike its endogenous form, is very unstable and catalyzes the oxidation of succinate by phenazine methosulphate or ferricyanide but not by cytochrome c or methylene blue. However, when soluble enzyme is re-incorporated within the particles of the heart-muscle preparation deprived of this enzyme, it becomes an integral part of the respiratory chain and re-acquires all the properties of its endogenous form, that is insolubility, stability and reactivity towards the cytochrome system and methylene blue. The succinic dehydrogenase-cytochrome system thus reconstituted, on addition of cytochrome c, actively catalyzes the oxidation of succinate by molecular oxygen. It shows normal oxidation and reduction of its cytochrome components and is susceptible to all inhibitors in the same way as is the succinic oxidase system of an untreated heart-muscle preparation. The comparative study of a soluble succinic dehydrogenase and a particulate succinic oxidase system revealed: (i) that pyrophosphate, like malonate and oxalacetate, acts as a competitive inhibitor of the succinic dehydrogenase itself; (ii) that narcotics have two sites of action: like antimycin they break the link between cytochromes b and c1 and, unlike antimycin, they also inhibit, but to a lesser degree, the activity of succinic dehydrogenase; and (iii) that cyanide can react with three components of this system in three different ways: a rapid and perfectly reversible reaction with cytochrome oxidase, a slow and still reversible reaction with oxidized soluble cytochrome c and a slow, irreversible reaction with the succinic dehydrogenase itself. It was previously shown (Tsou 1951) that the incubation of heart-muscle preparation with cyanide irreversibly inhibited the oxidation of succinate via the cytochrome system or methylene blue. Although cyanide treatment of soluble succinic dehydrogenase does not inhibit its catalytic activity in the oxidation of succinate by phenazine methosulphate, it prevents the dehydrogenase from anchoring itself within the particles of the heart-muscle preparation deprived of this enzyme, and so the reconstitution of the particulate succinic oxidase system does not take place. As the cyanide effect of Tsou is slow, irreversible, sensitive to temperature, complete and can be prevented by reducing substances, such as succinate acting as a hydrogen donor, or sodium dithionite, but not by malonate or pyrophosphate, we postulated: (i) that this effect is due to an irreversible reduction by cyanide of a disulphide group of succinic dehydrogenase to one thiol and one thiocyanate group, and (ii) th at this disulphide group originates by a reversible oxidation of a dithiol group which is present in addition to, and differs in several respects from the dithiol group of Hopkins.