When, however, a solution of a hexosephosphate is exposed to the action of either yeast-juice or zymin, entirely or partially freed from co-enzyme, this sugar, being no longer fermented, accumulates and can be examined. It has thus been found [Harden and Young, [1910, 2]] that a sugar is in fact produced in this way which can be fermented by living yeast and exhibits the reactions of fructose, although the presence of other hexoses is not excluded. The products of the enzymic hydrolysis of the hexosephosphates therefore appear to be the same as, or similar to, those formed by the action of acids [Young, [1909]].
A further consequence of these facts is that a hexosephosphate will yield carbon dioxide and alcohol when it is added to yeast-juice or zymin, and this has also been found to be the case [Harden and Young, [1910, 2]; Iwanoff, [1909, 1]]. [p057]
Mechanism of the Formation of Hexosediphosphoric Acid.
On this subject little is yet known, but a number of extremely interesting results, the interpretation of which is still doubtful, have been obtained by Euler and his colleagues. Euler has obtained a yeast [Yeast H of the St. Erik's brewery in Stockholm] which differs from Munich yeast in several respects. A maceration extract prepared from the yeast dried at 40° in a vacuum produces no effect on a glucose solution containing phosphate. If, however, the glucose solution be previously partially fermented with living yeast and then boiled and filtered, the addition of the extract prepared from Yeast H brings about the esterification of phosphoric acid without any accompanying evolution of carbon dioxide [Euler and Ohlsén, [1911], [1912]].
Euler interprets this as follows: (a) Glucose itself is not directly esterified, but must first undergo some preliminary change, which is brought about by the action of living yeast. No proof of the existence of a new modification of glucose in this solution has however been advanced, other than its behaviour to extract of Yeast H, so that Euler's conclusion cannot be unreservedly accepted. It is moreover possible and even more probable that some thermostable catalytic substance (perhaps a co-enzyme) passes from the yeast into the glucose solution and enables the yeast extract to attack the glucose and phosphoric acid. A very small degree of esterification was also produced when an extract having no action on glucose and phosphate was added to glucose which had been treated with 2 per cent. caustic soda for forty hours, but the nature of the compound formed was not ascertained [Euler and Johansson, [1912, 4]]. (b) The esterification of phosphoric acid without the evolution of carbon dioxide implies that the enzyme by which this process is effected is distinct from that which causes the actual decomposition of the sugar. Euler goes further than this and regards the enzyme as a purely synthetic one, giving it the name of hexosephosphatese to distinguish it from the hexosephosphatase which hydrolyses the hexosephosphate.
The evidence on which this conclusion is based cannot be regarded as satisfactory, inasmuch as it consists in the observation that in presence of sugar yeast extract does not hydrolyse the phosphoric ester. This, however, could not be expected since hydrolysis and synthesis under these conditions would ultimately proceed at equal rates.
In any case the adoption of this nomenclature is inconsistent with the conception of an enzyme as a catalyst and is therefore inadvisable until the reaction has been much more thoroughly studied. [p058]
It may further be pointed out that no proof has yet been advanced that the phosphoric ester produced without evolution of carbon dioxide is identical with hexosediphosphoric acid produced with evolution of carbon dioxide. It is by no means improbable that it represents some intermediate stage in the production of the latter (see p. [117]).
Euler's other results on this subject may be briefly summarised as follows:—
(1) In presence of excess of sugar the esterification of the phosphoric acid proceeds by a monomolecular reaction and is most rapid in faintly alkaline reaction [Euler and Kullberg, [1911, 3]].