8. REACTIONS OF INORGANIC SUBSTANCES
379
strated in green sulfur bacteria, particularly in Chlorobium limicola by
Kamen and Vernon (110) and in C. thiosulfatophilum by Gibson and
Larsen (111).
Several cytochromes are also present in the thiobacilli. Klimer et al.
(112) extracted single cytochrome s from T. thioparus. Despite its close
physiological similarity to this organism, Thiobacillus X has a different
cytochrome constitution, exhibiting several soluble cytochromes of type
c, as shown by Trudinger (113). One of these (553.5 ιτΐμ,) is autoxidizable, though its oxidation-reduction potential is strongly positive
(E' 0 = -f0.210 volt) and is reduced rapidly by thiosulfate at pH 7 in
the presence of the thiosulfate-oxidizing enzyme extracted from the
same bacteria. The subcellular particles which are sedimented at high
speeds contain other insoluble cytochrome compounds and a cytochrome oxidase; in the presence of the soluble cytochrome 553.5 and the
thiosulfate-oxidizing enzyme, the oxidation of thiosulfate can thus be
coupled with the consumption of molecular oxygen.
Aubert et al. (114) extracted and purified a soluble cytochrome c
from T. denitrificans which is not autoxidizable (E' 0 = +0.270 volt) and
constitutes a remarkably large proportion (3.5%) of the dry weight of the
bacteria. The same authors (115) showed that in whole cells this cytochrome takes part in the transport of electrons between the oxidation of
thiosulfate and the reduction of nitrate to N 2 . Moreover, they found
that cell-free extracts of T. denitrificans exhibit slight thiosulfate oxidase activity but contain a very active sulflte reductase. This is the
first bit of experimental evidence in support of an intermediary role of
sulflte in thiosulfate oxidation by thiobacilli. Like the sulfite oxidase of
dog liver, that of T. denitrificans requires hypoxanthine as a cofactor.
It is coupled to oxygen by cytochrome c and a cytochrome oxidase of
classical type, which is also present in extracts.
6. Phosphorylations
In discussing the mechanism of assimilation of C0 2 by chemo-lithotrophic bacteria in general, it was pointed out that the thiobacilli
couple sulfur oxidation and C0 2 incorporation energetically by way of
high-energy phosphate bonds. The formation of ATP from ADP and
inorganic phosphate by cells of T. denitrificans was demonstrated experimentally by Milhaud et al. (116) with the use of P
32 .
In sulfo-oxidizing bacteria, phosphorylations probably are involved
not only in energy transport but also in the intermediary metabolism of
the sulfur compounds themselves. The first observation along these lines
was made by Knaysi (117), who noted the formation of intracellular
vacuoles containing both sulfur and volutin (polymetaphosphates) dur-
379
strated in green sulfur bacteria, particularly in Chlorobium limicola by
Kamen and Vernon (110) and in C. thiosulfatophilum by Gibson and
Larsen (111).
Several cytochromes are also present in the thiobacilli. Klimer et al.
(112) extracted single cytochrome s from T. thioparus. Despite its close
physiological similarity to this organism, Thiobacillus X has a different
cytochrome constitution, exhibiting several soluble cytochromes of type
c, as shown by Trudinger (113). One of these (553.5 ιτΐμ,) is autoxidizable, though its oxidation-reduction potential is strongly positive
(E' 0 = -f0.210 volt) and is reduced rapidly by thiosulfate at pH 7 in
the presence of the thiosulfate-oxidizing enzyme extracted from the
same bacteria. The subcellular particles which are sedimented at high
speeds contain other insoluble cytochrome compounds and a cytochrome oxidase; in the presence of the soluble cytochrome 553.5 and the
thiosulfate-oxidizing enzyme, the oxidation of thiosulfate can thus be
coupled with the consumption of molecular oxygen.
Aubert et al. (114) extracted and purified a soluble cytochrome c
from T. denitrificans which is not autoxidizable (E' 0 = +0.270 volt) and
constitutes a remarkably large proportion (3.5%) of the dry weight of the
bacteria. The same authors (115) showed that in whole cells this cytochrome takes part in the transport of electrons between the oxidation of
thiosulfate and the reduction of nitrate to N 2 . Moreover, they found
that cell-free extracts of T. denitrificans exhibit slight thiosulfate oxidase activity but contain a very active sulflte reductase. This is the
first bit of experimental evidence in support of an intermediary role of
sulflte in thiosulfate oxidation by thiobacilli. Like the sulfite oxidase of
dog liver, that of T. denitrificans requires hypoxanthine as a cofactor.
It is coupled to oxygen by cytochrome c and a cytochrome oxidase of
classical type, which is also present in extracts.
6. Phosphorylations
In discussing the mechanism of assimilation of C0 2 by chemo-lithotrophic bacteria in general, it was pointed out that the thiobacilli
couple sulfur oxidation and C0 2 incorporation energetically by way of
high-energy phosphate bonds. The formation of ATP from ADP and
inorganic phosphate by cells of T. denitrificans was demonstrated experimentally by Milhaud et al. (116) with the use of P
32 .
In sulfo-oxidizing bacteria, phosphorylations probably are involved
not only in energy transport but also in the intermediary metabolism of
the sulfur compounds themselves. The first observation along these lines
was made by Knaysi (117), who noted the formation of intracellular
vacuoles containing both sulfur and volutin (polymetaphosphates) dur-