Amylose phosphorolysis

The glycogen phosphorolysis of phosphorylase can be reverted, which makes it possible to enzymatically polymerize amylose as well as hybrid structures with amylose as outlined in the following section. 

Arsenate similarly replaces phosphate in various phosphorolysis reactions, so that sucrose phosphorylase catalyzes the hydrolysis of sucrose in its presence (23), potato phosphorylase can hydrolyze amylose and amylopectin (24), nucleoside phosphorylase can hydrolyze inosine... 

One possible way to solve this problem is to combine another enzyme, like SP, which produces a-GlP for GP. Waldmann and colleagues reported the combined use of SP and GP for the production of amylose from sucrose (1986). In this system, SP catalyzes the phosphorolysis of sucrose to produce a-GlP and fructose, and the a-GlP is next used as a substrate of GP to produce amylose. An interesting feature of this SP-GP system is that Pi produced in the second GP reaction is used as a substrate for the first SP reaction. The cooperative action by the two phosphorylases proceeds continuously with a constant Pi concentration, without any inhibition caused by an accumulation of Pi. Based on this SP-GP system, we have now established the process to manufacture essentially linear amylose with controlled molecular size, by using thermostable variants of SP and GP (Yanase et al., 2007 Ohdan et al., 2007). 

We next tried to provide a-GlP, not from sucrose, but from cellobiose by using CBP. When CBP from Clostridium thermocellum (Kim et al., 2002) and GP were incubated with cellobiose in the presence of Pi and maltooligosac-chride primer, linear a-1,4 glucan was synthesized wth a yield (38.6 %) that was much lower than in the SP-GP method (Ohdan et al., 2007). To improve the yield of amylose, mutarotase and glucose oxidase were added to the initial reaction mixture with the expectation that they would remove the glucose derived from the CBP reaction and consequently shift the equilibrium state to phosphorolysis. The yield of amylose was increased to 64.8% (Ohdan et al., 2007). 

Although the synthesis of amylose by P-enzyme proceeds by a multichain mechanism, the reverse reaction shows some degree of single-chain action. The final products of phosphorolysis are a-D-glucosyl phosphate and maltopentaose. 

The phosphorylases are everywhere present in the biosphere. An enz5rme of this type (phosphorylase-a) has been obtained crystalline from rabbit muscle. In the presence of an excess of orthophosphate the reaction occurs in the direction of phosphorolysis, whilst in the absence of orthophosphate or in the presence of an excess of glucose-l-phosphate, the synthesis of amylose takes place. The enzyme is also called glucose-l-phosphate amylose-transglucosidase. 

Phosphorylases have been described for the polysaccharides, glycogen and amylose, and for sucrose. Phosphorylases from both muscle and plants are able to cleave the 1, 4 -qj linkages of glycogen and starch to yield glucose-1-phosphate. Sucrose phosphorylase is a separate enzyme obtained from bacteria which produces glucose-l-phosphate and fructose from sucrose. Phosphorolysis has also been involved in the cleavage of the pentosidic... 

Phosphorylases are key enzymes of carbohydrate metabolism and catalyze the phosphorolysis of glycosidic bonds in oligo- and polysaccharide substrates. This type of reaction afforded amylose via in vitro polymerization of D-glucosyl... 

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