Pullulanase

Pullulanase is a class of enzymes that can catalyze the hydrolysis of a-1,6-glycosidic bonds in starch and other polysaccharide compounds to form amylose. According to the differences in protein structure and the classification basis of the Glycoside Hydrolase family (GH), in general, pullulanase can be classified into two different glycoside hydrolase protein families: GH13 family and GH57 family. Most of the pullulanase belongs to the GH13 family, and only a few of the pullulanase belongs to the GH57 family. In terms of the spatial structure of the protein, the GH13 family of pullulanase has the typical (a/B) 8 barrels structure of this family. The GH57 family amylopectin has a (a/B) 7 barrels structure.

Figure 1. Protein structure of pullulanase.

Properties

Different sources of pullulanase have different specificity for the action of the substrate, mainly manifested in the ability to decompose various pullulan oligosaccharides. Different types of pullulanase have different reaction conditions, such as yeast pullulanase, aerogenes pullulanase and actinomycete pullulanase. The optimum pH is 6.2, 5.6-7.2 and 5.0, respectively, and the optimum temperature They are 20℃, 45-50℃ and 60℃ respectively. Metal ions have an effect on the activity of pullulanase, Ca²⁺ can improve the pH stability and thermal stability of isoamylase, while Hg²⁺, Cu²⁺, Fe³⁺, Al³⁺ and other inhibitory activities.

Source

Pullulanase exists widely in nature, and is distributed in almost all kinds of organisms. Except for a small part of pullulanase comes from animals, plants and other organisms, most of the pullulanase in nature exists in microorganisms. There are certain differences in a series of properties of pullulanase from different sources in amino acid sequence, substrate specificity, enzymatic properties, protein spatial structure and so on.

Applications

Pullulanase is widely used in industry. In the process of starch hydrolysis, adding pullulanase can increase the degradation rate of raw materials, shorten the reaction time, increase the conversion rate, and inhibit and reduce the production of other by-products. In the process of preparing high glucose syrup, because the glucoamylase cannot degrade a-1,6-glycosidic bonds, it often fails to achieve the expected DE value for production. Although the conversion rate can be increased by increasing the amount of saccharification enzyme or reducing the concentration of starch slurry, in addition to increasing the production cost, this will also produce by-product isomaltose, which will cause trouble for the subsequent process. Adding pullulanase to the saccharification enzyme in the saccharification stage for synergy, in addition to inhibiting and reducing the production of isomaltose, can also reduce the amount of saccharification enzyme, improve starch utilization, speed up the saccharification process, and save production time. Maltose has a sweet taste, low crystallinity and hygroscopicity, and low sweetness. Maltitol produced after the hydrogenation reaction of maltose has lower calories and can be used in special foods targeting specific disease groups instead of sugars. In the production of high-malt syrup, traditionally, a-amylase and exo-saccharification enzyme (ie, β-amylase) are used in combination to prepare high-malt syrup. However, since these two enzymes can only degrade a-1,4-glycosidic bonds, a large amount of dextrin is inevitably produced during the production process, resulting in the maltose content not exceeding 60%. In the traditional process of preparing high maltose syrup, by adding a certain amount of pullulanase, the yield of maltose can be as high as 90%.

Reference

1. Bender H.; et al. Pullulanase (an amylopectin and glycogen debranching enzyme) from Aerobacter aerogenes. Methods Enzymol. Methods in Enzymology. 1996, 8: 555–559.