Triosephosphate dehydrogenase and

In the course of reactions 1 to-4, a molecule of glucose has been transformed into a mixture of F—1,6—PP and two triosephosphates. Now occurs the first anaerobic oxido-reduction (see p. 142) in which, in the presence of triosephosphate dehydrogenase and its coenzyme DPN, an internal oxido-reduction takes place forming 1,3-diphosphoglyceric acid, a molecule containing an energy-rich acylphosphate bond. 

A second phase in the study of the fermentative systems of bacteria was the preparation of cell-free extracts capable of fermenting glucose. Thus, it was demonstrated in 1940 that a cell-free juice could be prepared from E. coli which contained triosephosphate dehydrogenase and aldolase and which, in the presence of arsenate, DPN, and fluoride, could degrade hexose diphosphate to phosphoglyceric acid. ... 

Reaction (6) is similar to that catalyzed by the D-glyceric dehydrogenase described by Stafford et al. (328) and reported to be widely distributed in green leaves. However, preparations of the carboxyla-tion enzyme contain little or no triosephosphate dehydrogenase and the stimulation by TPN (387), found with crude spinach extracts, is absent with the purified preparations. Glyceraldehyde 3 phosphate has been shown not to be a free intermediate in PGA formation from RuDP (386). 

Z F6P + ATP + 2 NADH + H+ <-> 2 glycerol-3-phosphate + ADP + 2 NAD+ where F6P is fructose-6-phosphate, FDP is fructose-1,6-diphosphate, DHA-P is dihydroxyacetone phosphate, TIM is triosephosphate isomerase, and GDH is glycerol-3-phosphate dehydrogenase. The oxidation of NADH is a measure of the 6-PFK activity and is determined photometrically (decrease of OD per minute) [4]. 

Another assay for phosphoffuctokinase involves converting the fructose 1,6-diphosphate to dihydroxyacetone phosphate and glyceraldehyde 3-phosphate with aldolase, equilibrating the triosephosphates with triosephosphate isomerase, and then measuring the production of NADH on the oxidation of the glyceraldehyde phosphate by glyceraldehyde 3-phosphate dehydrogenase. 

A period in the United States in 1939 as a Rockefeller Fellow in the Harvard laboratory of E. J. Cohn widened his experience of proteins and knowledge of their physical chemistry and led to his first crystallization of a muscle jirotein, a myogen from rabbit muscle which was later shown hy other workers to be identical with triosephosphate dehydrogenase. 

Triosephosphate Isomerase, Glyceraldehyde-3-Phosphate Dehydrogenase, and Phosphoglycerate Kinase. These enzymes catalyze the next three steps of glycolysis and are nearly-uniformly represented in all organisms. The... 

Zinc may be involved in the biosynthesis of tryptophan, a precursor of indole-3-acetic acid (lAA) (51). Zinc also participates in the metabolism of the plant as an activator of several enzymes. Zinc may act as an activator for some phosphate transferring enzymes such as hexose kinase or triosephosphate dehydrogenase. Zinc deficiency results in the accumulation of soluble nitrogen compunds such as amides and amino acids (52). Apparently zinc plays an important role in protein synthesis. 

TABLE 1. The distribution of marker enzyme activites in fractions obtained from pine protoplasts by differential centrifugation. The total activites (pmol (mg chi) h l) was 22 for PEPcarboxylase (PEPcase), 32 for fiimarase, and 69 for NADP triosephosphate dehydrogenase (NADP-TPD), respectively. The values are the means frcxn two separate fraction experiments. 

In the case of pyruvate kinase from cat muscle, too, the folding patterns of the domains could be compared with those of other proteins. Of the three domains in this protein (A, B, and C) similarities were noted between A and the structtire of triosephosphate isomerase and between C and the nucleotide binding region of lactate dehydrogenase. 

The AG value deduced from the PMF is corrected by replacing classical vibrational partition functions by their quantum homolog. Recrossing, tunnelling and non-classical reflection effects can be included in the transmission coefficient by various procedures. This ensemble-average variational transition state theory with multidimensional tunnelling (EA-VTST/MT) method was applied to proton and hydride transfers in various enzymes such as yeast enolase, liver alcohol dehydrogenase and triosephosphate isomerase. For a review, see ref. 3 and the chapter by J. Gao in this book. 

The cycle contains two oxidations, each coupled with TPN (and not DPN which in general is the coenzyme required in glycolysis). Glycolysis is inhibited by fluoride and iodoacetate or bromacetate, the first affecting enolase and the second triosephosphate-dehydrogenase. 

Many protein systems betides those discussed have been subjected to oxidative studies. Some of these are carbonic anbydrase (52), papain, and catheptic-like enzymes (53-57), invertase (58), succinic dehydrogenase (59), triosephosphate dehydrogenase (60), glycerol oxidase (61), and scarlet fever toxin (62). 

Strict specificity toward a coenzyme regardless of the source material. Alcohol dehydrogenase isolated from yeast and horse liver reacts with DPN but not at all with TPN. Triosephosphate dehydrogenase might have been cited as another example of strict DPN specificity except that current... 

In the discussions of triosephosphate, acetaldehyde, and a-keto acid dehydrogenases, it will be seen that there is a fairly close approach to an understanding of the energetic coupling in these oxidations. What is still a major mystery is the mechanism whereby the oxidation of the reduced pyridine nucleotides serves as the major energy source in most forms of aerobic metabolism. 

A coupled enzyme system can also be used. The glycer-aldehyde-3-phosphate formed is converted to dihydroxy-acetone phosphate by triosephosphate isomerase and this is followed by reduction by glycerol phosphate dehydrogenase. The oxidation of NADH in this last stage is measured spectrophotometrically. 

There are four enzymes in the glycolytic system which have more than usual interest from the standpoint of this chapter viz., triosephosphate dehydrogenase (T.D.), enolase, aldolase, and phos-phoglucoisomerase. The interaction of T.D. with its substrate (3-p-triose), DPN, and Pi may be represented as follows ... 

It has been known for sometime that in the oxidation of phospho-glyceraldehyde to phosphoglyceric acid, ADP is required. Closer inspection indicated that two independent enzyme systems are involved, one being triosephosphate dehydrogenase which catalyzes the formation of 1,3-diphosphoglyceric acid, and the other a specific enzyme responsible for the transfer of the phosphate of 1,3-diphosphoglyceric acid to ADP. Adenylic acid is inert except in the presence of adenylic kinase. 

The second type of a-glycerolphosphate dehydrogenase was first described by Green" as a particulate system which was coupled to cytochrome c. This particulate system has now been solubilized by treatment with sodium desoxycholate. On further purification the enzyme was separated from triosephosphate dehydrogenase, isomerase, and catalase activities and no longer was capable of reducing cytochrome c. It also did not react with DPN, TPN, or FAD, although it readily reduced suitable dyes. The reaction product was dihydroxyacetone phosphate. 

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