Glycolytic Sequence Research Articles

Action of foot-and-mouth disease virus and metabolic poisons on bovine kidney culture cells.

SummaryBovine kidney culture cells were found to be resistant to high concentrations of poisons of the glycolytic cycle, the citric acid cycle, and the cytochrome oxidation chain, as measured by respiration and cell loosening from the glass. Infection with FMDV had no influence on the results, and it may be that respiration of the cells is more dependent on other metabolic cycles. Iodo-acetate and fluoride, present in concentrations which caused large decreases in glycolysis, decreased virus production more than any other poisons. Malonate had an insignificant effect on virus production, while the larger effect of fluoroacetate was probably due to its action in removing viral precursors. The fact that maintenance of infected cells in an anaerobic environment did not affect virus production was a further indication that glycolytic sequences of the host cell are more essential to viral replication than oxidative sequences.

Some aspects of the carbohydrate metabolism of Entamoeba histolytical

Summary Activity of the following enzymes has been demonstrated in cell-free preparations of Entamoeba histolytica: (a) phosphohexoisomerase (b) phosphohexokinase (c) aldolase (d) glucose-6-phosphate dehydrogenase and (e) the enzyme system concerned in the Entner-Doudoroff pathway. Glyceraldehyde-3-phosphate dehydrogenase, α-glycerophosphate dehydrogenase and 6-phosphogluconate dehydrogenase could not be detected. Hexokinase could not be routinely demonstrated. While the presence of the enzymes phosphohexoisomerase, phosphohexokinase and aldolase would indicate that the glycolytic sequence is present from glucose-6-phosphate to triosephosphate, the absence of glyceraldehyde-3-phosphate dehydrogenase suggests that the steps of this sequence from triosephosphate to pyruvic acid are absent at least in our cell-free preparations. The hexosemonophosphate shunt would appear to be eliminated as a possible pathway of glucose metabolism by the absence of 6-phosphogluconate dehydrogenase. The Entner-Doudoroff scheme offers a pathway by which glucose maybe metabolized to glyceraldehyde-3-phosphate and pyruvate in E. histolytica.

Metabolism of Sarcina Lutea: II. Isotopic evaluation of the routes of glucose utilization

Sets of curves have been constructed which relate the degree of labelling of pyruvate molecules derived from the metabolism of glucose-1,2 and 6- 14C to the percentage participation of the hexose-monophosphate oxidative cycle in systems where this and the Embden-Meyerhof glycolytic sequence are the only routes of glucose catabolism in operation. The curves used in the final interpretation of the data assume that hexosemonophosphate formed in the cyclic operation of the hexosemonophosphate oxidative pathway is degraded by the two routes in the same proportion as the original glucose. The existence of a pool of hexosemonophosphate common to both pathways, which is inherent in this assumption, receives support from the good agreement obtained with the three different position-labelled glucose substrates. The method has been applied to the pyruvic acid produced from position-labelled glucose in the presence of arsenite by suspensions of lyophilized Sarcina lutea. Metabolism of glucose-U- 14C confirmed that there was no formation of pyruvate from endogenous sources but revealed a significant fixation of CO 2 in the carboxyl group of pyruvate. After correction for this CO 2 fixation, the results obtained indicate that glucose is metabolized aerobically by both the Embden-Meyerhof and hexosemonophosphate oxidative pathways in the approximate ratio of 70 to 30. Data on the assimilation of glucose-U- 14C and 1- 14C suggest that this ratio does not vary appreciably during metabolism. Thus, although Sarcina lutea is unable to utilize glucose anaerobically, Embden-Meyerhof glycolysis constitutes the major aerobic pathway.

Inhibition of phosphoglucose isomerase.

THE key carbohydrate intermediate, glucose 6-phosphate, which may follow several alternative metabolic pathways including the 6-phosphogluconate oxidative cycle and the glycolytic sequence, enters the latter route by the phosphoglucose isomerase reaction. This reaction, in which glucose 6-phosphate is reversibly converted into fructose 6-phosphate, could conceivably play a part in the regulation of carbohydrate metabolism, although it has seldom been considered from this point of view, doubtless because of the extremely high activity of the enzyme in most tissues and because the reaction has so far appeared to be very little affected by extraneous influences. The present evidence suggesting that the enzyme is strongly inhibited by 6-phosphogluconic acid, an intermediate in the direct oxidative pathway, may therefore be of interest in connexion with studies on the mechanism responsible for the partitioning of carbohydrate between the direct oxidative pathway and the glycolytic route.

The Direct Oxidation Pathway in Plant Respiration.

An important development in biochemistry has been the discovery of enzymes which can attack glucose-6-phosphate oxidatively to produce pentose phosphate and C02. The diversion of glucose-6phosphate by such a pathway has been variously called the hexose monophosphate shunt and the oxidative pathway and the names of several eminent research workers are linked with the elucidation of the reactions involved (reviewed by Horecker (9)). Recent work on a great variety of tissues, including higher plants (1, 2, 7, 8) has demonstrated that such systems are widespread. In view of this accumulating evidence, some workers have been led to consider that this metabolic byway may have importance in addition to its being a probable route to the pentoses in vivo, and the further elucidation of the reactions subsequent to pentose formation have lent color to this view. In particular, the recognition of the importance of sedoheptulose phosphate and its breakdown products has made possible the formulation of a cyclic sequence of events whose operation might of itself account for the total oxidation of glucose (9) and which would, at the least, produce metabolites which are also intermediates in the classical Embden-Meyerhof-Parnas (E.M.P.) glycolytic sequence. Although the details of the reactions remain to be clarified, the recent findings, to which experiments on higher plant tissues have contributed (1, 2, 7, 8), render the operation of the direct oxidation pathway in normal respiration quite plausible. There is ample evidence, of course, for the operation of the E.M.P. sequence of reactions in plant materials (11); at the present time, however, no evidence concerning the relative importance of the two alternatives in plant respiration has been offered. The present investigation was carried out to obtain evidence bearing on this point. Use was made of the fact that the method of glucose breakdown will determine the relative rates of release of its constituent carbon atoms. (For a discussion of the application of this principle see Bloom, Stetten, and Stetten (5) and Bloom and Stetten (6)). If the glucose molecule were broken down by glycolysis which includes equilibration at the tri?se level, two pyruvate molecules would be produced in which carbon atom 1 (C-l) and C-6 of the glucose would appear as the methyl carbons of the acid, C-2 and C-5 as the carbonyl carbons, and C-3 and C-4 as the carboxyl carbons. Carbons 1, 2, and 3 of the original glucose would, therefore, be indistinguishable respectively from 6, 5, and 4, and in the subsequent oxidative breakdown of the pyruvate, each member of a pair would appear in the respiratory C02 at the same rate as its partner. (Of the pairs, (C-3 and C-4) would be expected to appear first in the C02 followed by (C-2 and C-5) and then (C-l and C-6).) Thus, if comparable samples of tissue were respiring on glucose-1-C14 and glucose-6C14 respectively, the contribution of C14 to the C02 given off would be the same in each case. If, on the contrary, a glucose molecule was metabolized by way of the oxidative pathway, the C02 from the glucose1-C14 would be expected to be initially higher in C14 han that from the glucose-6-C14 since C-l of the glucose molecule is the first to be converted to C02. Provided that no assimilation of carbon residues containing different amounts of C-l and C-6 occurred, total yield of C1402 from the two glucose samples would be the same when oxidation was complete regardless of the path of breakdown. In the present experiments, it should be stressed, the time intervals were such that only a few percent of the supplied glucose was respired and the yields of C1402 are, therefore, regarded as initial as distinct from total. In these experiments, then, initial yields of C1402 from 2 glucose samples were compared, and the ratio, % radiochemical yield from glucose 6-C14/% radiochemical yield from glucose 1-C14 evaluated. Clearly, a ratio of near unity would indicate that schemes other than E.M.P. were playing a very minor part in glucose breakdown, whereas a ratio of less than unity would implicate the participation of the direct oxidation pathway.