Pullulanase Activity

Pullulanase exhibits highest activity at pH 5 and is stable at pH 5-6. It shows relatively higher activity when the temperature is around 50°C-60°C, and is stable at 30°C-60°C, but when the temperature is higher than 60°C, it will be rapidly inactivated [6]. 

The optimal pH of pullulanase is around 5.0 for the hydrolysis reaction, whereas the optimal pH of pullulanase for the reverse synthesis reaction is aroimd 4.0-4.5, as shown in Fig. 4.2. 

Temperature can affect the catalytic reaction activity. At a certain temperature range, the increasing of temperature increased the enzyme activity. However, further increasing of the temperature might destroy the conformation of the enzyme, and may result in a drop of the conversion rate. On the other hand, the solubility of j0-CD and maltose can be improved and the viscosity of the reaction system can be reduced by increasing the temperature. The optimal temperature of pullulanase is around 60 C-80°C [6]. 

At low substrate concentration, the hydrolysis reaction of pullulanase plays a dominant role. 

Only high substrate concentration is favorable for the reverse synthesis reaction of pullulanase (Fig. 4.3). But when the substrate concentration is higher than 80%, the conversion rate of Mal-jS-CD starts to decrease. It might be because the viscosity of the reaction system becomes too high, which is harmful for the enzymatic reaction, when increasing the substrate concentration to above 80% [6]. 

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Arnosti, C., and D. J. Repeta. 1994a. Extracellular enzyme activity in anaerobic bacterial cultures Evidence of pullulanase activity among mesophilic marine bacteria. Applied and Environmental Microbiology 60 840-846. 

Lipase (Microbial) Activity for Medium- and Long-Chain Fatty Acids, (S3)105 Lysozyme Activity, (S3)106 Maltogenic Amylase Activity, 804 Milk-Clotting Activity, 805 Pancreatin Activity, 805 Pepsin Activity, 807 Phospholipase A2 Activity, 808 Phytase Activity, 808 Plant Proteolytic Activity, 810 Proteolytic Activity, Bacterial (PC), 811 Proteolytic Activity, Fungal (HUT), 812 Proteolytic Activity, Fungal (SAP), 813 Pullulanase Activity, 814 Trypsin Activity, 814 Enzyme Assays, 786 Enzyme-Hydrolyzed (Source) Protein,... 

Pullulanase activity in a Klebsiella species, and which is of course an enzyme of starch catabolism, has been shown to be associated with the outer membrane of the cells. ... 

A pullulanase in E. coli appears to be bound to the cell wall its properties are very similar to those of a pullulanase from Klebsiella (Aerobacter) aerogenes. However, no pullulanase activity was detected in cultures of E. coli grown on maltose. Neither this nor forty other strains of E. coli grew on pullulan, whereas all the strains grew on maltose, and some on maltotriose. The synthesis of pullulanase by various strains of Klebsiella (Aerobacter) aerogenes using maltose both as an inducer and as a source of carbon has been studied." Conditions leading to the synthesis of pullulanase were reported. 

Extremely thermostable amylases and pullulanases, active above 100 °C and stable/active at acidic pH values wiU significantly improve the industrial starch bioconversion processes of liquefaction, saccharification, and isomerization. The lack of enzymes with these characteristics means that the bioconversion of starch to glucose and fructose must be carried out in a multistage process (step 1 pH 6.0 to 6.5, 95 to 105 °C step 2 pH 4.5, 60 to 62 °C step 3 pH 7.0 to 8.5, 55 to 60 °C),... 

Carbohydrase (Bacillus acidopullulyticus) Produced as an off white to brown, amorphous powder or a liquid by controlled fermentation using Bacillus acidopullulyticus. Soluble in water (the solution is usually light yellow to dark brown), but practically insoluble in alcohol, in chloroform, and in ether. Major active principle pullulanase. Typical applications used in the hydrolysis of amylopectins and other branched polysaccharides. 

Calculations One pullulanase unit (PUN) is the amount of activity that under the conditions of the test, will liberate reducing sugars equivalent to 1 pimol of glucose per min. Determine the linear regression line for absorbance versus two times the glucose concentration (pug/mL) in the standards. Use the slope, I, in the following equation to determine the activity in the enzyme preparation ... 

One important point to be remembered when pullulanase is being used is its heat stability. Treatment of digests at 100° for a short time is not sufficient to render the enzyme inactive activity returns spontaneously on standing. This is more noticeable at pH 7 than at pH 5. It is most impor-... 

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