Lab Activity 30

Digestive Enzymes

Portland Community CollegeBI 233

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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

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Energy of Activation (EA)

• For a reaction to occur, an energy barrier must be overcome

• Enzymes make the energy barrier smaller

EAwithout enzyme

EA with enzyme

energyreleasedby the

reaction

products

starting substance

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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

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Enzymes

• Environmental conditions affect enzymes:• Temperature• pH• Salt concentration

• When you “denature” an enzyme, you change its shape

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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

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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

Substrate

Enzyme

Active site

NORMAL BINDING OF SUBSTRATE

Competitiveinhibitor

Non-competitiveinhibitor

ENZYME INHIBITION

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Condensation (aka Dehydration Synthesis)

• Two molecules combine• Water is a byproduct

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43

3

2

2

1

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Hydrolysis

• Type of cleavage reaction• Opposite of condensation

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1

2

2 3

3

4

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Chemical Digestion

• Most digestive enzymes catalyze hydrolysis reactions.

• Addition of H2O breaks polymers into smaller subunits (monomers, dimers ect..)

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Four types of Macromolecules

CarbohydratesProteins

Nucleic acids

monosaccharidesamino acidsfatty acids and glycerolnucleotides

polysaccharides

Monomer(s)

polypeptides

Lipids

Polymer(s)Class

fats, steroids phospholipids

polynucleotides

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Carbohydrate Digestion

• Goal #1: Break complex carbs (starch) down to oligosaccharides, trisaccharides, disaccharides

1. Salivary Amylase: (minor): breaks complex carbs (starch, glycogen) to oligosaccharides, trisaccharides, and disaccharides. Inactivated by gastric acid.

2. Pancreatic amylase: (major)3. Amylase in breast milk

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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

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Lugol’s IKI

• IKI: potassium iodide• Indicator for starch

• Turns black in the presence of starch

IKI alone Positive result for starch

Negative result for starch

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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)

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Benedict’s Solution

Before heating

After heating

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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

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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

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Protein Digestion

1. Brush-border membrane peptidases

2. Brush-border membrane amino acid transporters

3. Brush-border membrane di- and tripeptides transporters

4. Intracellular peptidases5. Basolateral-membrane amino acid

carriers6. Basolateral membrane di- and

tripeptides carriers

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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

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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

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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

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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

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The End