Demonstration glucose isomerase

The successful conversion of D-glucose into D-fructose on the industrial scale with immobilized D-glucose isomerase was a brilliant demonstration of the value of this kind of approach. Then followed a huge technical literature on enzyme immobilization, reviewed in Ref. 9 (page 353). We shall here restrict ourselves to the methods which have been utilized in the syntheses outlined in Tables II to X. We suggest to readers interested in theses techniques that they first use these methods. If they prove unsatisfactory, as there is a plethora of alternatives, other techniques, described in Refs. 8-10, may be tried a majority of readily available carbohydrate enzymes have been immobilized, often in several different ways. 

The first large-scale commercial application of cross-linked enzyme crystals was the use of glucose isomerase CLCs to produce high-fructose com syrup. While this is not a pharmaceutical or a biotechnological application, it is included here because it serves to demonstrate the economic viability of the technology in a very cost-sensitive business. In this application the CLCs were attached to the surface of a polystyrene-cellulose-titanium oxide composite carrier in a ratio of 9 1 carrier enzyme. The catalyst had a half-life of 150 days at 57°C, and 12-18 tons of dry sugar product could be produced per kilogram of enzyme [37],... 

The work reported here demonstrates that the microporous plastic sheet (MPS )is a viable support for glucose isomerase. 

D-Glucose Isomerases (o-Xylose isomerases).—A discussion of enzyme chemistry in the D-glucose isomerase field demonstrates its role in the displacement of enzyme equilibria by oxyanions (including germanate) to obtain high yields of D-fructose from D-glucose. ... 

A true glucose isomerase that did not require arsenate for its activity was discovered in Lactobacillus brevis [12]. Such enzymes were later also found in the genus Streptomyces and this source was first reported in 1965 [13]. Subsequent studies demonstrated that the Streptomyces enzyme responsible for this conversion is inducible and is rather a D-xylose isomerase, as the value oiK ... 

D-Xylose isomerases have been found in a wide range of microorganisms. As mentioned in Sect. 2.1, their substrate specificities are not confined to o-xylose (18) and D-glucose (17) but can include d- and L-arabinose (13 and 24) [17,74], D-ribose (20) [15,20,75], and L-rhamnose (6-deoxy-L-mannose, 26) [17], as well as D-allose (22) [17]. Very recent investigations already mentioned [74] have shown that L-ribose (40) as well as D-lyxose (41) are also accepted as substrates by some enzymes of the type under consideration. This would mean that of all eight aldopentoses, only L-lyxose and L-xylose have not yet been identified as substrates of D-xylose isomerases, demonstrating an intrinsic tolerance of these enzymes towards unusual substrates. 

Subsequently, isomerases from Aerobacter aerogenes strains which also converted L-xylose (110, Scheme 33) were purified [91,92] and the inducible enzymes from two other E. coli strains were compared [93]. It was demonstrated that both preparations were tetramers with masses around 350 kDa and were active with L-fucose (15) and D-arabinose (13). However, this was not the case for the other aldopentoses and hexoses probed such as L-arabinose (24), l-rhamnose (26), D-xylose (18), D-ribose (20), d- (17) or L-glucose (58), d- (44) or L-mannose (111), D-galactose (57) or D-fucose (6-deoxy-D-galactose, 112). Both enzymes were inhibited by alditols related to their natural substrates, exhibited their optimal catalytic activities in alkaline media, and were stimulated by the presence of Mn as well as Co but strongly inhibited by Cd. ... 

Roseman et al. (1958) purified the enzyme from microbial and mammalian extracts and demonstrated that fructose-6-phosphate, and not glucose-6-phosphate, was the required substrate. Ghosh et al. (1969) purified the enzyme from Neurospora crassa, E. coli, and rat liver until it was free from phosphoglucose isomerase and glutaminase. Using this more purified enzyme preparation, fructose-6-phosphate was shown to be the required substrate, and a 1 1 stoichiometry was demonstrated for glutamine and fructose-6-phosphate. 

To summarize these observations First, the enzymes of this segment of the photosynthetic carbon reduction cycle are not controlled co-ordinately a single inductive step does not appear to affect the activity (and presumably the production) of these three enzymes in the same way. Second, the kinds of control mechanisms which appear to occur here are (1) a direct and prompt effect of illumination, reflected in the rapid increase in ribulose-diphosphate carboxylase activity, and (2) a more indirect kind of control, possibly involving induction of the other enzymes (e.g., ribose-phosphate isomerase) by small molecules produced in photosynthesis. Unfortunately, it has not been possible to demonstrate an increase in the level of either the isomerase or kinase by administration of glucose and some other carbohydrates to etiolated leaves in darkness. 

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