D-Glucan Phosphorylase

0 X 10 ) in the presence of sodium dodecyl sulphate or guanidinium hydrochloride, contains pyridoxal phosphate, and is stable in the pH-range 

The amino-acid composition of the enzyme was also determined. 

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Cell extracts of Cryptococcus laurentii, a yeast which synthesizes amylose when grown at low pH, contain an a-D-glucan phosphorylase (EC 2.4.1.1) which has been characterized. Evidence for a possible role of this enzyme in the biosynthesis of amylose was reported. 

In a procedure described for the simultaneous large-scale isolation of pullulanase and (1 4)-oc-D-glucan phosphorylase from Klebsiella pneumoniae, the pullulanase is solubilized from the cell wall by cholate treatment cells and cell debris are removed by partition in a poly(ethyleneglycol)-dextran two-phase system and from the upper phase of this system the pullulanase is isolated by ultrafiltration and precipitation with A -cetyl-iV, A -trimethylammonium bromide to 80% purity with 70% yield and in an a-amylase-free state. 

The fact that hydroxy-groups at positions C-2, C-3, C-4, or C-6 in the D-glucose moiety of a-D-glucopyranosyl phosphate are not essential for its substrate specificity in the transglycosylation reaction with a-(l 4)-D-glucan phosphorylase in plant polysaccharides was probably an important observation. Recent discoveries on the distribution of side chains in plant polysaccharides... 

The synthesis of linear 4 —> 1-a-D-glucans from D-glucopyranosyl phosphate by the action of phosphorylases has been shown by comparison of results of methylation and end-group assay and viscosity determination,209 and by potentiometric, iodine titrations82 on the product. The chain length of the enzymic product (100 to 200 D-glucose units) is less than that of the natural component. Whether this is due to impure enzymes cannot yet... 

In the field of polymer science, the most extensively used transferase is phosphorylase (systematic name (1 4)-a-D-glucan phosphate a-D-glucosyltransferase EC 2.4.1.1). Although this enzyme is responsible for the depolymerization of linear a-( 1 4) glycosidic chains in vivo it can also be used to synthesize linear a-( 1 4) glycosidic chains (amylose) in vitro. 

Sivak. M. N., 1992. (1— 4)-a-D-Glucan synthesis by a chloroplastic phosphorylase isolated from spinach leaves is independent of added primer. Carbohyd. Res. 227, 241-255. 

Glucose 1-phosphate can also be formed directly by an hexosyltrans-ferase, phosphorylase (1,4-a-D-glucan orthophosphate a-glucosyltrans-ferase EC 2.4.1.1)... 

In a model experiment, glucoamylase (ex Rhizopus niveus) was shown to inhibit a-D-glucan synthesis as catalysed by potato phosphorylase (see p. 498).29 ... 

For a glucan to be a substrate for starch phosphorylase, it must be longer than maltotetraose. Shorter oligosaccharides, however, can be used by glucosyltransferases such as D-enzyme (EC 2.4.1.25) in the reaction... 

Fig. 2 Main reactions catalyzed by sucrose-utilizing transglucosidases. The synthesis of a-glucans (Glc) linked through either a-1,6 a-1,3, a-1,4, or a-1,2 osidic linkages depending on the enzyme specificity is obtained by glucansucrases. The formation of a-D-glucose-1-phosphate from orthophosphate is obtained using sucrose-phosphorylases. The mechanism involves a double-inversion at the anomeric center via the formation of a covalent P-D-glucosyl enzyme intermediate...

One property of phosphorylase that has not been widely used, but that could often be advantageous, is its ability to degrade or-n-glucans, releasing D-glucose instead of D-glucosyl phosphate when the reaction is conducted in the presence of arsenate rather than phosphate. It is considered that D-glucosyl arsenate is formed initially, but that it is labile... 

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