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Plants with high starch content such as corn, potatoes, rice, sorghum, wheat, and cassava have played significant roles in human development. From hamanch (manioc beer) to high fructose corn syrup, both low tech and high tech approaches to the use of starch prevail. Starch molecules are glucose polymers linked together by the alpha-1,4 and alpha-1,6 glycosidic bonds. A family of enzymes known as the glycosidases aid in the breakdown of starch and other glucose polymers such as celluslose and glycogen to smaller sugars, and finally, to glucose. All of these enzymes hydrolyze glycosidic bonds, but some are also multifunctional. Sequence data for many of the glycosidases are well described in terms of their functional roles (active sites and protein folding) and a great deal of research can be found on evolutionary relationships between these enzymes in different taxa. Not only alpha-amylases, but also beta-amylases and starch debranching enzymes such as the pullulanases and glucosidases also contain the beta/alpha barrel domain including catalytic amino acids. The mechanisms differ, but the relatedness of these enzymes is clear. Amino acid sequences of the beta-strands are well conserved within this family of enzymes. This provides a rationale for using the glycosidases to investigate the evolutionary relationships between organisms. Glycosidases and the modification of corn starch In the commercial production of maltodextrins and corn syrups, starch is hydrolyzed using an alpha-amylase either alone or combined with other enzymes.
What does a glycosidase look like? The alpha-amylases contain eight alpha-helices and eight beta-strands in beta alpha/beta alpha order. The alpha-helices provide rigidity to the catalytic sites and substrate binding sites which are contained within the beta-strands.
How does a glycosidase break down starch? If we look at the alpha-amylase enzyme, we can find both the catalytic sites and the substrate binding site. The amino acid sequence of alpha-amylases may vary, but there are specific aspartic acid and glutamic acid units found in the beta-strand region that are responsible for the catalysis of glycosidic bond cleavage. Other amino acid units such as histidine are necessary for enzyme activity involved in establishing conformation and binding of the substrate. Steps in enzymatic hydrolysis of starch
Conservation of active sites and catalytic sites Not only alpha-amylases, but also beta-amylases and starch debranching enzymes such as the pullulanases and isoamylases also contain the beta/alpha barrel domain including the same catalytic amino acids. The mechanisms differ, but the relatedness of these enzymes is clear. Amino acid sequences of the beta-strands are well conserved within this family of enzymes. This provides a rationale for using the glycosidases to investigate the evolutionary relationships between organisms.
Solving the problem of synonyms Besides providing essential information on enzyme classification, E.C. numbers are very useful for doing searches when variations of enzyme names are encountered. Enzyme functions are classified by E.C. numbers:
Listed below are the E.C. numbers of several starch hydrolyzing enzymes further characterized by their mode of action:
VisualizationStructural data files for many of the glycosidases are readily available from the Protein Data Bank. All pdb files have unique 4 character names that include numbers and letters. The Protein Data Bank Education page provides a good introduction to the international repository for 3-D molecular structure data. See: http://www.rcsb.org/pdb/education.html
Data Amylase FASTA Sequences Tools PDB Viewers, RasMol program and VRML browser Biology Workbench ConSurf Server PyMOL RSCB Protein Data Bank Summaries and structural analyses of PDB data files Protein Explorer
In addition to introductory Microbes Count! Activities, the following scenarios are presented: Scenario 1. Glycosidases and the modification of corn starch …In the commercial production of maltodextrins and corn syrups, starch is hydrolyzed using an alpha-amylase either alone or combined with other enzymes.
Scenario 2. Enzyme replacement therapy: Should you try increasing your own levels of alpha-amylase? There are a number of over-the-counter products that contain enzymes that aid in the digestion of proteins, starches, fats and dairy foods. For example, Lactaid® contains the enzyme lactase for helping the digestion of dairy products.
The product contains the following enzymes:
Scenario 3. Alpha-amylase inhibitors, weight loss, and beans When trying to lose weight, dieters limit the amount of starch in their diet. The usual amounts of starchy foods, such as potatoes, bread, beans, corn and pasta, are reduced. Starch provides from 500 to 700 calories per day in the average American adult diet. A gram of starch, when digested and absorbed, provides 4 calories. Individuals may consume as much as 1,500 or more calories per day from starch contained in their foods. However, starch is a large molecule that cannot be absorbed if it is not first broken down. Undigested starch will pass on through the digestive tract. Plant extracts containing AIU's (alpha-amylase inhibiting units) such as phaseotein from legumes are said to inhibit the absorption of starch by blocking the enzyme alpha-amylase.
Scenario 4. Allergic to your breakfast cereal? Is the alpha-amylase inhibitor in wheat the culprit? Using serum samples collected from children with a known wheat allergy and one adult with baker's asthma, a wheat protein was identified which bound IgE. Control serum samples were collected from wheat-tolerant patients. No IgE binding to this wheat protein was demonstrated in any of the control subjects. Samples representing the 15 kd wheat protein (isoelective point 5.85) were selected and the N-terminal peptide sequence of this protein (residues 1 to 20) matched to a wheat alpha-amylase inhibitor.
Scenario 5. Investigating the human alpha-glucosidase gene Glucose is a major source of energy for the body. It is stored in the form of glycogen in both the liver and muscles and later released with the help of enzymes. Persons affected by glycogen storage disease (GSD) have an inherited defect in one of the enzymes responsible for forming or releasing glycogen as it is needed by the body during exercise and/or between meals. There are eleven types of GSD known at this time. Read the following brochure written by a mother whose son inherited an infantile form of Pompe's Disease which reduces glycogen storage function to less than 2% of normal. This is an autosomal recessive disorder that is always fatal.
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Clinical Genetics Site NiceZyme View of ENZYME: EC 3.2.1.3 GSD II Database: A register of mutations in Human acid alpha-glucosidase Looking into Glycosidases: A Bioinformatics Resource for Biology Students Starch: Chemistry and Technology. Whistler, BeMiller & Paschall, Eds. 1984. San Diego : Academic Press. Starch: Properties and Potential, Galliard, Ed. 1987. New York : John Wiley & Sons.Note: Names used for this disease:
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Author: Ethel Stanley 2005
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