1. Enzymes
Packet #13
Chapter #8
Friday, November 23, 2018

2. Properties of Enzymes Catalytic Efficiency Specificity
Enzymes catalyze (speed up) reactions 103 to 106 faster than uncatalyzed reactions
Lower the activation energy
Using the chemical equation model, enzymes work in only one direction—as they will not catalyze a reverse reaction
Specificity
Enzymes are very specific
Interacting with one, or few, specific substrates and catalyzing only one type of chemical reaction
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3. Properties of Enzymes II
Cofactors
Some enzymes associate with a nonprotein cofactor that is needed for enzymic activity…
Zn2+
Fe2+
…and with organic molecules that are often derivatives of vitamins
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4. Properties of Enzymes III
Location within the cell
Many enzymes are localized in specific organelles within the cell
Allows isolation of substrate or product from other competing reactions
Provides a favorable environment for the reaction
Allows organization of the 1000’s of enzymes present in the cell into purposeful pathways.
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5. Naming Enzymes
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6. Naming of Enzymes Most historically Substrate + ase
Sucrase
Catalase
Mallerase
International Union Biochemistry and Molecular Biology
4 digit Nomenclature Committee Numbering System
1st
Major Class of Activity
Only six classes recognized
2nd
Subclass
Type of bond acted on
3rd
Group acted upon
Cofactor required
4th
Serial Number
Sequence order
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7. Classes of Enzymes
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8. Classes of Enzymes I Oxidoreductases Transferases Hydrolases Lyases
Catalyze oxidation-reduction reactions
Transferases
Catalyze transfer of C, N or P containing groups
Hydrolases
Catalyze cleavage of bonds by addition of water
Catalyze hydrolysis reactions
Lyases
Catalyze cleavage of C-C, C-S and certain C-N bonds
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9. Classes of Enzymes II Isomerases Ligases
Catalyze conversion of a molecule from one isomeric form to another
Catalyze racemization of optical or geometric isomers
Catalyze isomerization
Change from one isomer to another
Ligases
Catalyze certain reactions in which two molecules join in a process coupled to the hydrolysis of ATP.
Catalyze formation of bonds between carbon and O, S, N coupled with hydrolysis of high energy phosphates (ATP)
Condensation of 2 substrates with splitting of ATP
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10. Enzyme Structure
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11. Structure Enzymes are active proteins.
On the structure of an enzyme, one would find an active site where the chemical reaction takes place.
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12. Structure II
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13. Function & Specificity
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14. How Enzymes Work
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15. Gibb’s Free Energy Free Energy
The portion of a system’s energy that can perform work when temperature and pressure are uniform throughout the system.
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16. Reading Energy Charts
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17. Exergonic Reaction
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18. Activation Energy Activation Energy Transition State
The energy difference between reactants and the transition state
Determines how rapidly the reaction occurs at a given temperature
The lower the activation energy, the faster the reaction will occur
The higher the activation energy, the slower the reaction will occur
Transition State
Represents the highest-energy structure involved in the process of a chemical reaction
A chemical reaction must have enough energy to overcome the “transition state.”
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19. Energy Charts
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20. Factors that Change the Shape of Enzymes
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21. Changing Shape of Enzyme I
Temperature
Increases the kinetic motion
Breaks the hydrogen bonds
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22. Changing Shape of Enzyme IpH
Changes the ionic charges
Alters the shape
If the pH becomes basic, the acidic amino acid side chains will lose H+ ions
If the pH becomes acidic, the basic amino acid side chains will gain H+ ions
Causes the ionic bonds, that help stabilize the tertiary structures of proteins, to break. Resulting in the denaturation of the enzyme.
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23. Inhibiting Enzyme Function
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24. Inhibiting Enzyme Function
Inhibitors
Chemicals that binds to enzyme and changes its activity
Competitive
Inhibitor that binds at the active site
Non-competitive
Inhibitor that binds at alternate site other than active site
Poisons
Organo-phosphorous compounds
Insecticides
Bind to enzymes of the nervous system and kills the organism
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25. Inhibitors
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26. Enzyme Kinetics
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27. Enzyme Kinetics Way of describing properties of enzymes
Mathematical
Graphical expression
Expression of reaction rates of enzymes
A  B + C
Please read Chapter 8
Section #4
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28. Graphical Curves of Enzyme Activity
Rate vs. Enzyme
Ml substrate/min Rate
Rate vs. pH
Reveals the optimum pH
Rate vs. Temperature
Reveals the optimum temperature
Rate vs. Substrate
Shows a saturation curve
Most definitive curve of enzyme activity
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29. Michaelis-Menten Enzyme Curve
Michaelus and Menten proposed a simple model that accounts for most of the features of enzyme-catalyzed reactions.
In this model, the enzyme reversibly combines with its substrate to form an Enzyme-Substrate Complex that subsequently breaks down to product.
Results in the regeneration of a free enzyme.
E + S ↔ ES  E + P
S = substrate
E = Enzyme
ES = Enzyme-substrate complex
K1, k-1, k2 = rate constants
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30. Michaelis-Menten Equation
Describes how reaction velocity varies with substrate concentration
Rate (Reaction Velocity) vs. Substrate Concentration
V0 = Vmax [S]/Km + [S]
V0 = initial reaction velocity
Vmax = maximal velocity
Km = Michaelis constant = (k-1 + k2)/k1
Is the substrate concentration at which rate is one-half the maximal velocity
A measure of affinity of enzyme for a substrate
[S] = Substrate Concentration
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31. Michaelis-Menten Enzyme Curve
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32. Michaelis-Menten Equation
Assumptions (3)
The concentration of substrate is greater than the concentration of enzymes
Remember, only one substrate is able to bind at the active site of an enzyme at any time.
The rate of formation of the enzyme-substrate complex is equal to the breakdown of the enzyme-substrate complex
To either
E + S
E + P
Recall equation from earlier slide.
Initial velocity
Only used in the analysis of enzyme reactions
Meaning, the rate of reaction is measured as soon as enzyme and substrate are mixed
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33. Conclusions about Michaelis-Menten Kinetics
Characteristics of Km
Km = substrate level at ½ Vmax
Does not vary with the concentration of enzyme
Small Km
Reflects high affinity(an attraction to or liking for something) of the enzyme for substrate
Why?
Because a low concentration of substrate is needed to reach a velocity of ½ Vmax
Large Km
Reflects low affinity of the enzyme for substrate
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34. Conclusions about Michaelis-Menten Kinetics
Relationship of Velocity to Enzyme Concentration
Rate of the reaction is directly proportional to the enzyme concentration at all substrate concentrations
Example
If the enzyme concentration is halved, the initial rate of the reaction (v0) is reduced to one half that of the original
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35. Conclusions about Michaelis-Menten Kinetics
Order of Reaction
Recall from Chemistry
Will leave the details of this conclusion out.
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36. Lineweaver-Burke Plot
When the reaction velocity is plotted against the substrate concentration, it is not always possible to determine when Vmax has been achieved.
Due to the gradual upward slope of the hyperbolic curve at high substrate concentration.
However, if 1/V0 is plotted vs 1/[S] , a straight line is obtained.
This plot is known as the Lineweaver-Burke Plot
Can be used to calculate Km
Vmax
Determines the mechanism of action of enzyme inhibitors
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37. Lineweaver-Burke Equation
1/V0 = Km/Vmax[s] + 1/Vmax
The intercept on the x axis
-1/Km
The intercept on the y axis
1/Vmax
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38. Michaelis-Menten & Lineweaver-Burke
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39. The Effect of Inhibitors
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40. Competitive Inhibition
Effect on Vmax
Vmax is the same in the presence of a competitive inhibitor
Effect on Km
Michaelis constant, Km, is increased in the presence of a competitive inhibitor
Effect of Lineweaver-Burke Plot
Vmax is unchanged
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41. Competitive Inhibition
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42. Succinate Dehydrogenase
Enzyme that catalyzes the oxidation of succinate to fumarate during cell Respiration
Malonate
Structurally similar to the substrate succinate
Binds at the active site of the enzyme
Results in an increase of the substrate succinate in the cell
However, the probability of the active site being occupied by the substrate, instead of the inhibitor, increases
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43. Non-Competitive Inhibition
Effect on Vmax
Vmax is decreased
Cannot overcome by increasing the amount of substrate
Effect on Km
Michaelis constant, Km, is the same
Non-competitive inhibitors do not interfere with the binding of substrate to enzyme
Effect of Lineweaver-Burke Plot
Vmax decreases
Km is unchanged
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44. Effects of Inhibitors
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45. Lead Poisoning & Lactam Antibiotics
Lead forms covalent bonds with the sulfhydryl side chains of cysteine in proteins
The binding of the heavy metal shows non-competitive inhibition
Drugs an behave as enzyme inhibitors
Lactam antibiotics
Penicillin
Amoxicillin
Inhibit one or more enzymes of bacteria walls
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46. Metabolic Pathways
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47. Regulation of Enzyme Activity
The regulation of the reaction velocity of enzymes is essential if the organism is to coordinate its numerous metabolic pathways—The control of an organism’s metabolism.
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48. Feedback Inhibition
An end product inhibits an initial pathway enzyme by altering efficiency of enzyme action
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49. Allosteric Regulation
Results in changes an enzymes shape and function by binding to an allosteric site
Specific receptor site on some part of the enzyme molecule remote from the active site
Allosteric inhibitor, binds at the allosteric site, and stabilizes the inactive form of the enzyme
Makes the enzyme non-functional
Activator, also binds at the allosteric site, and stabilizes the active form on the enzyme
Makes the enzyme functional
ATP and ADP are examples
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50. Allosteric Regulation
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