Lab Activity 30

1. Lab Activity 30 Digestive Enzymes Portland Community College BI 233

2. Cellular Reactions • All molecules have energy barriers to prevent spontaneous breakdown • Enzymes speed up the cell’s chemical reactions by lowering energy barriers • The amount of energy needed to overcome the energy barrier is the energy of activation (EA) • Enzymes lower the EA for chemical reactions to begin; decreases the amount of energy the reactants must absorb

3. Energy of Activation (EA) • For a reaction to occur, an energy barrier must be overcome • Enzymes make the energy barrier smaller EA without enzyme EA with enzyme starting substance energy released by the reaction products

4. Enzymes • A protein that acts as biological catalysts by lowering the activation energy • Increase the rate of chemical reactions • Are highly specific, they only act on one substrate or reaction • Not consumed in the reaction E + S ES complex  E + P *If there is no enzyme, the reaction will still happen, eventually… Enzymes cannot make a non-spontaneous reaction spontaneous

5. Enzymes • Environmental conditions affect enzymes: • Temperature • pH • Salt concentration • When you “denature” an enzyme, you change its shape

6. Enzyme Helpers • Some enzymes require non-protein cofactors • Some are inorganic metal ions of zinc, iron, and other trace elements • Some are organic molecules called coenzymes • Includes vitamins or altered vitamin components

7. Enzyme Inhibitors • Inhibitors block enzyme action • A competitive inhibitor takes the place of a substrate in the active site • A noncompetitive inhibitor alters an enzyme’s function by changing its shape NORMAL BINDING OF SUBSTRATE Substrate Active site Enzyme Non-competitiveinhibitor Competitiveinhibitor ENZYME INHIBITION

8. Condensation (aka Dehydration Synthesis) • Two molecules combine • Water is a byproduct 1 2 3 1 2 3 4

9. Hydrolysis • Type of cleavage reaction • Opposite of condensation 1 2 3 4 2 1 3

10. Chemical Digestion • Most digestive enzymes catalyze hydrolysis reactions. • Addition of H2O breaks polymers into smaller subunits (monomers, dimers ect..)

11. Four types of Macromolecules Monomer(s) Polymer(s) Class Carbohydrates monosaccharides polysaccharides Proteins amino acids polypeptides fats, steroids phospholipids fatty acids and glycerol Lipids Nucleic acids nucleotides polynucleotides

12. Carbohydrate Digestion • Goal #1: Break complex carbs (starch) down to oligosaccharides, trisaccharides, disaccharides • Salivary Amylase: (minor): breaks complex carbs (starch, glycogen) to oligosaccharides, trisaccharides, and disaccharides. Inactivated by gastric acid. • Pancreatic amylase: (major) • Amylase in breast milk

13. Carbohydrate Digestion • Goal #2: further breakdown into monosaccharides • Use brush border enzymes on microvilli of small intestine • 1. Lactase: breaks lactose into glucose + galactose • 2. Maltase: breaks maltose into 2 glucoses, (also works on oligosaccharides) • 3. Sucrase: breaks sucrose into glucose + fructose

14. Lugol’s IKI • IKI: potassium iodide • Indicator for starch • Turns black in the presence of starch Positive result for starch Negative result for starch IKI alone

15. Benedict’s Solution • Benedict's solution is used to detect the sugars glucose or maltose • It is a blue solution that will turn red-orange (brick red) when heated in the presence of glucose or maltose • (note that a sucrose solution would not change color)

16. Benedict’s Solution Before heating After heating

17. Protein Digestion • Goal #1: Break Proteins down into smaller polypeptides • Pepsin (pepsinogen from stomach’s chief cells) • HCL in stomach denatures the proteins to enhance digestion. • Pancreatic enzymes: trypsin, elastase, chymotrypsin & carboxypeptidase: break large polypeptides to small polypeptides & peptides

18. Protein Digestion • Goal #2: break polypeptides amino acids. • On brush border: peptidases • Inside cytoplasm of intestinal cells: several dipeptidases, tripeptidase break absorbed dipeptides and tripeptides into amino acids

19. Protein Digestion • Brush-border membrane peptidases • Brush-border membrane amino acid transporters • Brush-border membrane di- and tripeptides transporters • Intracellular peptidases • Basolateral-membrane amino acid carriers • Basolateral membrane di- and tripeptides carriers

20. Protein Digestion • BAPNA is a dye attached to an amino acid via a peptide bond. • Peptide bonds are bonds that link amino acids in polypeptides. • When the peptide bond is broken in BAPNA with trypsin, the dye is released and turns yellow

21. Fat Digestion • Goal #1: Break Big fat droplets into smaller droplets • Bile salts emulsify • Smaller spheres of fat have higher surface/volume • Makes lipase (water soluble enzyme that can’t penetrate fat droplet) more efficient

22. Fat Digestion • Goal #2: Break triglycerides into monoglycerides and fatty acids • Gastric Lipase from chief cells in fundus of stomach (20% of digestion) • Digestion products: monoglycerides and fatty acids • Pancreatic Lipase (80%), • Digestion products: monoglycerides and fatty acids • Milk-derived lipase: in breast milk • Digestion products: fatty acids and glycerol

23. Litmus Cream • Litmus is a pH indicator- purple in storage bottle • It comes mixed with cream (a triglyceride source) • Triglyceride digestion by lipase releases fatty acids. • These fatty acids drop the pH, and litmus turns PINK

24. The End The End