Maltotetraose amylase

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... 

Using the pure anomers of maltose in the same way, we found that crystalline hog pancreatic a-amylase causes the very rapid synthesis of maltotetraose from a-maltose but not from /3-maltose whereas, crystalline sweet potato /3-amylase causes the very rapid synthesis of the same compound, specifically from /3-maltose. Configurational inversion again marks this latter condensation as glycosyl-hydrogen interchange, or glycosyl transfer. 

Maltose and maltotriose were synthesized from a-glucosyl fluoride by all the a-amylases. Maltotetraose and sizeable amounts of the pentaose... 

Hasegawa, K., Kubota, M., and Matsuura, Y. (1999). Role of catalytic residues in a-amylases as evidenced by the structures of the product-complexed mutants of a maltotetraose-forming amylase. Protein Eng., 12, 819-824. 

In these a-amylase reactions, glucose is not the primary product. Glucose is formed as a secondary product in the hydrolysis of maltotriose, maltotetraose and branched dextrins. Using 14C-reducing end labeled maltodextrins, G3 to G8, Robyt and French19 showed that porcine pancreatic a-amylase has different specificities for hydrolyzing the different linkages of the maltodextrins. The different frequencies for... 

Uchida et al.220b reported the synthesis of 6m-deoxy maltodextrins by B. macerans CGTase transglycosylation reactions between mono-6-O-p-toluene sulfonyl cyclomal-tohexaose and maltose. The best inhibitors for human salivary a-amylase and human pancreatic a-amylase were 6m-deoxy maltotetraose and 6m-deoxy maltopentaose. 

In a similar use of a-amylase, Parrish and Whelan249 treated potato starch with crystalline human salivary a-amylase and obtained a phosphorylated maltotetraose that had previously been reported by Postemak250 and that was the smallest phospho-dextrin formed. They determined its structure to be 63-phosphomaItotetraose, similar in structure to the smallest a-limit dextrin, 63-a- D-glucopyranosylmaltotriose, formed by this enzyme and porcine pancreatic a-amylase. 

Figure 9.72 Chromatograms of the action patterns of maltoheptaose after the indicated periods of incubation with a-amylase and a-glucosidase. Peaks 1, glucose 2, maltose 3, maltotriose 4, maltotetraose 5, maltopentaose (x) compound A 6, maltohexaose 7, maltoheptaose. (A) Pure maltoheptaose used for the assay. (B) Blank sample before the addition of substrate. (C-H) Chromatograms after 1, 5, 10,15, 20, and 30 minutes, respectively, of incubation. Chromatographic conditions column, 10 jum Nucleosil SA (250 mm X 4 mm) solvent, acetonitrile-water (72.527.5) flow rate, 0.7 mL/min temperature, 27°C detection, differential refractometer, full scale = 2 X 10-6 refractive index units. (From Haegel et aL, 1981.)...

The enzyme is readily prepared, in an apparently pure form, from the culture filtrates of this pseudomonad, and has a specific activity comparable to that of crystalline, sweet-potato befa-amylase. The exo action of the enzyme is analogous to that of heta-amylase, but maltotetraose residues, not maltose residues, are removed. The action of the enzyme is halted by branch points, so that limit dextrins are produced from glycogen and amylopectin. The specificity of the enzyme for linkages in the region of branch points has not yet been ascertained. Some applications of this enzyme will be discussed later (see p. 318). 

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. 

Fig. 17.—Effect of Prior Treatment with aZpha-Amylase on the Products Liberated by Fullulanase from the Maltotetraose Residues in Pullulan.

The other tetrasaccharide component (Rg 0.26) had the same Rg value as that of maltotetraose, which was identified by methylation and the action of beta amylase. 

Thus, the variability of the actions of several a-amylases from different origins on elsinan is consistent with the conventional classification of amylases. The enzymes having high activities on the relatively lower maltosaccharides, such as maltotetraose and maltopentaose, are likely to hydrolyze elsinan. On the other hand, the incapability of certain a-amylases, e. g., the liquefying type amylases, may be due to their affinities to higher maltosaccharides. This can be supported by the fact that Taka amylase shows a lower activity to yield particular tetrasaccharide and heptasaccharide. 

The main degradation product (75-90%) resulting from the action of selected a-amylases on the a-D-glucan of Elsinoe leucospila has been identified as the trisaccharide O-a-D-glucopyranosyl-(l- 3)-0 -D-glucopyranosyl-(1 4)-D-glucose. This confirms previous reports that the polysaccharide consists chiefly of malto-triose units and a small proportion of maltotetraose units joined by (1 3)-a-D-... 

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