Cellobiose phosphorylase

Langsford et al. reported that Cellulomonas fimi culture supernatants contained cellulase and proteinase activities, for which there appeared to be a relationship. Glucose repressed the synthesis of both activities and cellulose induced both 60), Adding cellulose to Cellulomonas sp. (NRCC 2406) cultures stimulated growth and improved production of cellulases 61). Optimum conditions for growth and cellulase production were pH 6.5 and 30 C. The addition of glucose in the presence of cellulose inhibited growth. Several species of Cellulomonas have cellobiose phosphorylase. 

CELLOBIOSE PHOSPHORYLASE CHORISMATE SYNTHASE COBALAMIN ADENOSYLTRANSFERASE 3-DEHYDROQUINATE SYNTHASE... 

Recently Kuriki and coworkers succeeded in producing glucose-1-phosphate in situ during the enzymatic polymerization of amylose. By using sucrose phosphorylase or cellobiose phosphorylase, the monomer was produced during the polymerization from inorganic phosphate and sucrose or cellobiose, respectively [119-121]. 

Cellobiose formation by the cellobiose phosphorylase reversible reaction. 

Phospholylases, as summarized in Table 28.1, catalyze the reversible phospho-rolysis of polysaccharides or oligosaccharides, and produce phosphorylated mono-saccharides. Among such enzymes a-glucan phosphorylase (GP, EC 2.4.1.1), sucrose phosphorylase (SP, EC 2.4.1.7) and cellobiose phosphorylase (CBP, EC 2.4.1.20) are of great interest, since they can produce a-glucose 1-phosphate (a-GlP) from three major biomasses starch, sucrose, and cellulose. Only these three are described in this paper, a comprehensive review of other phosphorylases can be found in Kitaoka and Hayashi (2002). 

Cellobiose Phosphorylase CBP catalyzes the reversible phos-phorolysis of cellobiose into a-GIP and glucose as shown in equation 28.3 ... 

Kim, Y. K., Kitaoka, M., Krishnareddy, M., Mori, Y., and Hayashi, K. 2002. Kinetic studies of a recombinant cellobiose phosphorylase (CBP) of the Clostridium thermocellum YM4 strain expressed in Escherichia coli. J. Biochem., 132, 197-203. 

Kitaoka, M., Sasaki, T., and Taniguchi, H. 1992. Conversion of sucrose into cellobiose using sucrose phosphorylase, xylose isomerase and cellobiose phosphorylase. Denpun Kagaku, 39, 281-283. 

It is probably as stable probes of the specificity of enzymes that carba-sugars find their justification. A striking example of their utility is illustrated by the elucidation of the stereochemical selectivity of the reverse reaction of cellobiose phosphorylase, where only the 5a-carba-(3-D-glucopyranose was a substrate of the reaction — not the a anomer (Scheme 8.2) [11]. 

Kitaoka, M, Ogawa, S, Taniguchi, H, A cellobiose phosphorylase from cellvibrio-gilvus recognizes only the beta-D-form of 5a-carba-glucopyranose, Carbohydr. Res., 247, 355-359, 1993. 

Specific labelling of cellobiose can be readily achieved by use of cellobiose phosphorylase (24). 

The combination of the oligosaccharide glucotransferase with the hydrolase and the cellobiose phosphorylase, then accounts fully for the conservation of the glucosyl bond energy predicted by the growth experiments. 

The only other enzymes which could yield glucose from dimer and tetramer are the phosphorylases (cellobiose phosphorylase, etc.). 

The presence of cellobiose phosphorylase (EC 2.4.l.a) catalyzing the reversible phosphorolysis of cellobiose to a-D-glucose-1-phosphate and glucose was first demonstrated by Ayers (3, 4) in cell-free extracts of R. flavefaciens. Recently, Tyler and Leatherwood (60) reported on the presence of an epimerase catalyzing the conversion of cellobiose into glucosylmannose in the culture filtrate of R. albus. 

Cellobiose must be cleaved into its constituent monosaccharides in order to be metabolized by . coli. There are two main ways in which this reaction can occur, hydrolysis and phosphorolysis. P-Glucosidase and cellobiose phosphorylase from Saccharophagus degradans were expressed in E. coli. The results showed that phosphorolysis cells tolerate common inhibitors (sodium acetate) more effectively and produce recombinant proteins more effectively than hydrolysis cells. However, hydrolysis cells utilize xylose more effectively in combination with cellobiose [192]. 

Similarly, the choice of enzymes in pathway design is of importance. In the case of the conversion of cellulose to starch, cellobiose phosphorylase and a-glucan phosphorylase are responsible for reversibly eonverting from eello-biose to amylose or vice versa. It was found that a-gluean phosphorylase from potato is a key enzyme to drive the reaction toward starch synthesis. In contrast, the same enzyme from Clostridium thermocellum eannot generate amylase from cellobiose because it prefers the starch degradation direetion. ... 

Nidetzky, B., Griessler, R., Schwarz, A., Splechtna, B., 2004. Cellobiose phosphorylase from Cellulomonas uda gene cloning and expression in Escherichia coli, and application of the recombinant enzyme in a glycosynthase-type reaction. Journal of Molecular Catalysis B Enzymatic 29, 241—248. 

Reichenbecher, M., Lottspeich, E., Bronnenmeier, K., 1997. Piuification and properties of a cellobiose phosphorylase (CepA) and a cellodextrin phosphorylase (CepB) from the cellulolytic thermophile Clostridium stercorarium. European Joiunal of Biochemistry 247,262—267. 

Figure 4.6 Coproduction of synthetic amylose and ethanol from cellulose via multi-enzymes and an ethanol-producing yeast. EG endoglucanase, CBH cellobiohydrolase, CBP cellobiose phosphorylase, PGP potato a-glucan phosphorylase.

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