Pullulan Pullulanase

Pullulan is hydrolyzed by pullulanase at the a-(l - 6) bonds, producing maltotriose plus some maltotetraose. Salivary alpha amylase cleaves at the maltotetraosyl units, when the a-(1 -> 4) linkage next to the a-(1 -> 6) bond and towards the reducing end of the maltotetraose unit is split (dotted arrow marked A in 108). The size of units released by alpha amylase, as judged... 

Isoamylase [9067-73-6] (glycogen-6-glucanohydrolase) and pullulanase [9012-47-9] (pullulan-6-glucanohydrolase) hydrolyzes a-l,6-glucosidic bonds of starch. When amylopectin is treated with a pullulanase, linear amylose fragments are obtained. Using a heat- and acid-stable pullulanase in combination with saccharifying enzymes makes the starch conversion reactions more efficient (71). 

The plant and bacterial enzymes capable of hydrolyzing pullulan do not have identical specificities. In particular, the plant enzymes have little or no action on glycogen and phytoglycogen under conditions in which they readily hydrolyze amylopectin and its /3-dextrin. To stress this difference (the bacterial enzymes are capable of degrading both glycogen and phytoglycogen), Manners (1997) recommended different nomenclature for bacterial enzymes, to be called pullulanase, and the plant enzymes, to be called limit dextrinases. 

Pullulanase is an extracellular enzyme of Aerobacter aerogenes that causes essentially quantitative hydrolysis of pullulan to maltotriose. The enzyme is readily prepared in a crude form that is free from other carbohydrases, and is important in structural studies because it debranches amylopectin and glycogen. When the A. aerogenes is grown in continuous culture, the enzyme is bound to the cells, but it can be released by detergents, and purified by ion-exchange chromatography. This purified enzyme has been crystallized. ... 

Pullulan, fungal, 372, 382 Pullulanase, 360, 373 purification by chromatography, 283 Pyran-2,6-dimethanol, tetrahydro-, 62 Pyran-2-methanol, tetrahydro-, 62 Pyran-3-methanol, tetrahydro-, 62 Pyranosides, history, 3 Pyrazole, 3-(D-eri/f/iro-l,2,3-trihydroxypro-pyl)-, 163... 

Fig. 3-118. Gradient analysis of pullulan after enzymatic digestion with pullulanase. - Chromatographic conditions see Fig. 3-115 injection 50 pL of a 0.2% solution.

In contrast to the previously described pullulanases, pullulan hydrolases types I and II are unable to hydrolyze a-l,6-glycosidic linkages in pullulan or in branched... 

Pullulanase type I hydrolyzes a-1,6 linkages in amylopectin, pullulan or limit dextrins with high specificity. Pullulan is completely degraded in a random fashion... 

Pullulanase Type I pullulan, branched polysaccharides [endoacting a-1,6] maltotriose, linear oligosaccharides Klebsiella pneumoniae Bacillus acydopullulyticus Fervidobacterium pennivorans Thermotoga maritima... 

Alpha-dextrin endo-1,6-alpha-glucosidase. Pullulanase. Pullulan 6-glucanohydrolase. Limit dextrinase. Debranching enzyme. 3.2.1.41 Starch-debranching enzyme, hydrolyses (l-6)-alpha-glucosidic linkages in pullulan and starch to form maltotriose. 

Modified substrates may be used in testing the action patterns of purified enzymes. Thus, periodate-oxidized laminaran was used by Smith and coworkers to show the exo action of Basidiomycete (l- 3)-j8-n-glucanase. The oxidized-amylose substrate described earlier (see p. 266) has been used with alpha- and heta-amylases to illustrate how the method is completely general, and applicable to all classes of polysaccharides and poIysaccharidases. ° By use of oxidized pullulan, the endo nature of pullulanase action was confirmed. ... 

The discovery, shortly thereafter, of the enzyme pullulanase from the bacterium Aerobacter aerogenes helped considerably in further structural analysis of the polysaccharide. Hydrolysis of pullulan by diis enzyme showed that essentially no products containing (l->6)-ff-i>-giucosidic linkages were formed, the preponderant product being maltotriose. It was thus clear that the enzyme hydrolyzes (l->6)-a-D-glucosidic link-... 

A more careful examination of the products of action of pullulanase on pullulan showed the presence of a tetrasaccharide, in addition to maltotriose. In the sample examined, the tetrasaccharide was estimated to constitute 7% of the polysaccharide. The complete hydrolysis of this tetrasaccharide to maltose by sweet-potato befo-amylase, as well as optical rotation measurements, characterized it as maltotetraose. As earlier work had suggested that pullulan is a linear molecule, it was, therefore, of interest to ascertain how the maltotetraose units are arranged in the molecule. The two most likely possibilities are illustrated in Fig. 16b,c. 

Pullulanase Type II (glucoamylase EC3.2.1.3) Hydrolysis of a-l, 4-glycosidic bond from the non-reducing end of pullulan to generate glucose. 

Pullulanase Type I (Pullulanase EC3.2.1.41) Hydrolysis of a-l, 6-glycosidic bond in pullulan and straight-chain oligosaccharides to generate maltotriose and linear oligosaccharides. 

Pullulanase Type I (neopullulanase) Hydrolysis of a-1, 4-glycosidic bond in pullulan to generate Pan sugar. 

An a-amylase found in the culture filtrate of a strain of Thermoactinomyces vulgaris was purified by ammonium sulphate fractionation, and ion-exchange chromatographies.The purified enzyme showed a single band on disc gel electrophoresis. The pH optimum, temperature optimum, and isoelectric point were pH 5.0, 70 C, and pi 5.2, respectively. The a-amylase was stabilized by Ca and was found to hydrolyse pullulan to panose. Therefore, the hydrolytic pattern of this enzyme is different from those of pullulanase and isopullulanase. 

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