Starch Hydrolysis by Amylase

We thus elucidated that three of the four cellulase components are endo- or random-type and the other is exo-type. However, it is difficult to distinguish between the components of least or lowest random-type and those of exo-type. It is rather easy to identify an endo-type cellulase component. In contrast, it is very difficult to determine a cellulase to be exo-type because if the enzyme has a glycosyl-transferring activity the hydrolysis product is not a single sort, which is one of the necessary conditions to be an exo-type. Based on our experiments, measurement of the time course of CMC using a sample of medium substitution degree seems to be the best method of diagnosis to determine a cellulase component to be endo- or exo-type. With some enzymes, direction of mutarotation of reaction products is useful to resolve this problem, as is illustrated by the classic example of the starch hydrolysis by a- and /3-amylases. If this is true for our cellulases, the mutarotation of reaction products would be a... 

The (3-amylases catalyze starch hydrolysis by a mechanism that gives inversion of configuration at the anomeric center. All known (3-amylases have an exo-mechanism and act on the non-reducing ends of starch polymer chains or starch polymer-derived chains. There are two general classes of (3-amylases, those that are classically known as (3-amylases and produce (3-maltose, and those that are known as glucoamylases and produce (3-D-glucose. 

For starch hydrolysis by microbial a-amylase, the following activation energies, which lie between the limits stated in section 2.5.4.2, were derived from e. g. the Arrhenius diagram (Fig. 2.35) ... 

Unlike many of the catalysts that chemists use in the laboratory, enzymes are usually specific in their action. Often, in tact, an enzyme will catalyze only a single reaction of a single compound, called the enzyme s substrate. For example, the enzyme amylase, found in the human digestive tract, catalyzes only the hydrolysis of starch to yield glucose cellulose and other polysaccharides are untouched by amylase. 

Maltose Digestion by amylase or hydrolysis of starch. Germinating cereals and malt. ... 

Table X 4 summarizes similar data for the hydrolysis by maltase-free malt alpha amylase of beta dextrins obtained from arrowroot starch by the action of beta amylase. The beta dextrins were precipitated with alcohol from the reaction mixture of arrowroot starch after it had reached a limit in the hydrolysis at 60% theoretical maltose. The beta dextrins were hydrolyzed extensively by malt alpha amylase. Glucose was liberated in very small amounts even in the later stages of the hydrolysis of these beta dextrins maltose was liberated in appreciable amounts and, at equivalent hydrolyses, appeared to be formed somewhat more rapidly from the beta dextrins (Table X) than from the untreated starch (Table IX). Upon hydrolysis with malt alpha amylase the molecular weights of the beta dextrins dropped appreciably but not as extensively as when arrowroot starch was hydrolyzed directly by malt alpha amylase.

More specific hydrolysis may be achieved by the use of enzymes. Thus, the enzyme a-amylase in saliva and in the gut is able to catalyse hydrolysis of al 4 bonds throughout the starch molecule to give mainly maltose, with some glucose and maltotriose, the trisaccharide of glucose. Amylose is hydrolysed completely by this enzyme, but the al 6 bonds of amylopectin are not affected. Another digestive enzyme, a-l,6-glucosidase, is required for this reaction. Finally, pancreatic maltase completes the hydrolysis by hydrolysing maltose and maltotriose. 

The investigations carried out by Professor French and his students were based on sound experimental approaches and on intuitive theoretical considerations. The latter often resulted in new experiments for testing a hypothesis. On the basis of theoretical considerations, Professor French proposed a model for the structure of the amylopectin molecule, and the distribution of the linear chains in this molecule. This model was tested by utilizing enzymes that selectively cleave the linear chains, and the results substantiated the theoretical deductions. He proposed a theory on the nature and types of reactions occurring in the formation of the enzyme - starch complex during the hydrolysis of starch by amylases. In this theory, the idea of multiple attack per single encounter of enzyme with substrate was advanced. The theory has been supported by results from several types of experiments on the hydrolysis of starch with human salivary and porcine pancreatic amylases. The rates of formation of products, and the nature of the products of the action of amylase on starch, were determined at reaction conditions of unfavorable pH, elevated temperatures, and increased viscosity. The nature of the products was found to be dramatically affected by the conditions utilized for the enzymic hydrolysis, and could be accounted for by the theory of the multiple attack per single encounter of substrate and enzyme. 

Resistant starch escapes hydrolysis by starch-specific enzymes (see Commentary). Studies have shown that resistant starch, upon treatment with DMSO, becomes solubilized and, thus, hydrolyzable by amylase enzymes. 

Hanes, C. S. Studies on Plant Amylases The Effect of Starch Concentration upon the Velocity of Hydrolysis by the Amylase of Germinated Barley. Biochem. J. 1932, 26, 1406-1421. 

A typical profile for starch hydrolysis using soluble amylase is shown in Fig. 1. The profile predicted by the kinetics model is also shown. Clearly, the model describes the experimental concentration profiles very well. The model curves also show the insensitivity of the model fit to Km there is very little difference in the quality of the model predictions with Km = 5 g/L and Km = 50 g/L. Similarly, a typical profile for starch hydrolysis using immobilized amylase is shown in Fig. 2. The model also predicts these data very well, with little sensitivity to the Km value. 

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