Ribulose-5-phosphate formation

Fig. 5. A simplified metabolic scheme of ethanol formation from glucose and xylose. Enzyme abbreviations GPDH Glucose 6-phosphate 1-dehydrogenase, PGDH Phosphogluconate dehydrogenase, PGI Glucose 6-phosphate-isomerase, RKI Ribose 5-phosphate isomerase, RPE Ribulose phosphate 3-epimerase, TAL Transaldolase, TKL Transketolase, XDH Xylitol dehydrogenase, XK-. Xylulokinase, XR Xylose reductase...

The oxidative portion of the pentose phosphate pathway consists of three reactions that lead to the formation of ribulose 5-phosphate, C02, and two molecules of NADPH for each molecule of glucose 6-phosphate oxidized (Figure 13.2). This portion of the pathway is particularly important in the liver and lactating mammary glands, which are active in the biosynthesis of fatty acids, in the adrenal cortex, which is active in the NADPH-dependent synthesis of steroids, and in erythrocytes, which require NADPH to keep glutathione reduced. 

Phosphate and bicarbonate ions are important substrates for many enzymatic processes and as such have regulatory functions. Bicarbonate controls the key enzyme of photosynthesis, ribulose bisphosphate carboxylase, by carbamate formation (Fig. 13-12). Chloride ions activate amylases and may affect the action of "G proteins" that mediate hormone actions. Other observed effects of ions are too numerous to mention. 

In bacteria, which lack formate dehydrogenase, formaldehyde can be oxidized to C02 to provide energy beginning with the reactions of Eq. 17-51. The resulting fructose 6-P is isomerized to glucose 6-P, which is then dehydrogenated via Eq. 17-12 to form C02 and the regenerating substrate ribulose 5-phosphate. 

Photosynthesis. The formation of carbohydrates in green plants by the process of photosynthesis is described in ihc entry on Photosynthesis. The synthetic mechanism involves the addition of carbon dioxide to ribulose-1,5-diphosphate and the subsequent formation of two molecules of 3-phosphoglyccric acid which are reduced to glyceraldehyde-3-phosphate. The triose phosphates are utilized to again from ribulose-5-phosphates by enzymes of the pentose phosphate cycle Phosphorylation or ribulose-5-phosphate with ATP regenerates ribulose-1.5-diphosphate to accept another molecule of carbon dioxide. See also Phosphorylation (Photosynthetlc). 

D-Arabinose-5-phosphate isomerase, the first key enzyme in the synthesis of KDO, catalyzes the interconversion of D-ribulose-5-phosphate and D-arabinose-5-phosphate. This enzyme was briefly studied by Volk (18) and later by Lim and Cohen (19). Due to the instability of the enzyme, we have only purified this enzyme 100-fold. The reversible reaction is readily monitored by measuring the formation of the keto-sugar from the aldo-sugar by the method of Dische and Borenfreund (20). The K values for D-ribulose-5-phojsphate and D-arabinose-5-phosphate are 0.9 to 1.5 and 1 to 3 x 10 M, respectively. 

Measurements of the crude specific activity (mmoles of product synthesized per minute per mg of protein in the supernatant after a 50,000 x g centrifugation) of the two isomerases in E. coli indicated that the conversion of D-ribulose-5-phosphate to D-ri-bose-5-phosphate was approximately 20- to 30-fold greater than the conversion of D-ribulose-5-phosphate to D-arabinose-5-phosphate. This rate of reaction strongly pulls the reaction substrate to D-ribose-5-phosphate, since the isomerase reaction at equilibrium strongly favors the formation of the aldo-sugar over the key intermediate D-ribulose-5-phosphate. 

The precursors for riboflavin biosynthesis in plants and microorganisms are guanosine triphosphate and ribulose 5-phosphate. As shown in Figure 7.3, the first step is hydrolytic opening of the imidazole ring of GTP, with release of carbon-8 as formate, and concomitant release of pyrophosphate. This is the same as the first reaction in the synthesis ofpterins (Section 10.2.4), but utilizes a different isoenzyme of GTP cyclohydrolase (Bacher et al., 2000, 2001). 

Regeneration of ribulose bisphosphate occurs by the same reactions that occur in the hexose-monophosphate shunt. The hexose monophosphate shunt interconverts 3-, 4-, 5-, 6- and 7-carbon sugar phosphates. In the formation of one molecule of glucose from CO2, carrying out the previous reactions six times is necessary. Making the six molecules of ribulose-bisphosphate required for fixing six COi s occurs by the following sets of reactions ... 

Figure 5 Formation of the pyridoxine ring in vitamin B5. (A) deoxyxylulose phosphate-dependent pathway (B) deoxyxylulose phosphate-independent pathway. 43, 1-deoxy-D-xylulose 5-phosphate 47, 3-amlno-1-hydroxyacetone 1-phosphate 48, pyridoxine 5 -phosphate 49, ribulose 5-phosphate 50, dihydroxyacetone phosphate 39, pyridoxal 5 -phosphate.

A third ketopentulose ester, xlulose 5-phosphate (Xlu 5-P) Fig- 2, was isolated as a product of Rib 5-P metabolism by Ashwell and Hickman (34). It was also shown by Srere et al. (35) that Xlu 5-P, rather than the earlier assigned Ru 5-P, was a definitive substrate of TK. Ribulose 5-phosphate-3 -epimerase (Fig. 2) catalyzed the formation of Xlu 5-P and imparted the transconflguration to the hydroxyls at carbons 3 and 4, which is a necessary stereochemical condition for substrate reactivity with TK. The 3 -epimerase was purified from a bacterial source... 

The pentose phosphate pathway also catalyzes the interconversion of three-, four-, five-, six-, and seven-carbon sugars in a series of non-oxidative reactions. All these reactions occur in the cytosol, and in plants part of the pentose phosphate pathway also participates in the formation of hexoses from CO2 in photosynthesis. Thus, D-ribulose 5-phosphate can be directly converted into D-ribose 5-phosphate by phosphopentose isomerase, or to D-xylulose 5-phosphate by phosphopentose epimerase. D-Xylulose 5-phosphate can then be combined with D-ribose 5-phosphate to give rise to sedoheptulose 7-phosphate and glyceraldehyde-3-phosphate. This reaction is a transfer of a two-carbon unit catalyzed by transketolase. Both products of this reaction can be further converted into erythrose 4-phosphate and fructose 6-phosphate. The four-carbon sugar phosphate erythrose 4-phosphate can then enter into another transketolase-catalyzed reaction with the D-xylulose 5-phosphate to form glyceraldehyde 3-phosphate and fructose 6-phosphate, both of which can finally enter glycolysis. 

An enzymic counterpart of these complex base-catalysed rearrangements of sugars may be the reaction catalysed by 4-phospho-3,4-dihydroxy-2-butanone synthetase. The enzyme catalyses the formation of the eponymous intermediate in secondary metabolism from ribulose 5-phosphate. Labelling studies indicated that C1-C3 of the substrate became C1-C3 of the product, that H3 of the substrate derived from solvent and that C4 was lost as formate. X-ray crystal structures of the native enzyme and a partly active mutant in complex with the substrate are available. The active site of the enzyme from Met ha-nococcus jannaschii contains two metals, which can be any divalent cations of the approximate radius of Mg " or Mn ", the two usually observed. Their disposition is very reminiscent of those in the hydride transfer aldose-ketose isomerases, but also to ribulose-5-phosphate carboxylase, which works by an enolisation mechanism, so the enolisation route suggested by Steinbacher et al. is repeated in Figure 6.14, as is the Bilik-type alkyl group shift, for which an equivalent reverse aldol-aldol mechanism cannot be written. 

Fig. 3. Binding curve of a DNA aptamer that is selected to bind a protein enzyme known as Bcs1. Bcs1 is a CDP-ribitol synthase (a bifunctional enzyme that catalyzes two consecutive chemical reactions leading to the formation of CDP-ribitol the reduction of ribulose 5-phosphate using NADPH, and cytidylyltransfer using CTP) from Haemophilus influenzae.JM is the corresponding CDP-ribitol synthase from Staphylococcus aureus. Radioactively labeled DNA aptamer (—0.1 nM) was incubated with 0, 25, 50, 75,100, and 200 nM of each enzyme in the selection buffer and the mixtures were analyzed using 8% nondenaturing polyacrylamide gel. The fraction of radioactivity in the shifted DNA band is calculated and plotted against the protein concentration.

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