Enzymes: Overview

• Functional proteins that catalyse biological reactions
• Involved in all essential body reactions
• Found in all body tissues
– Seen in serum following cellular injury or from degraded cells
• Decrease the amount of free energy needed to activate a specific reaction

• Not altered or consumed during reaction
• Reusable
• Accelerate speed of reactions

• Holoenzyme
– Functional unit
– Consists of:
• Apoenzyme
• Cofactor/coenzyme
• Proenzyme/zymogen
– Inactive enzyme
Holoenzyme

• Role
– Increase reaction rates while not being consumed or altered
Enzyme
– Substrate Product

• Active site
– Specific area of the enzyme structure that participates in the reaction(s)/interacts with the substrate

• Allosteric site
– Non-active site
– May interact with other substances resulting in overall enzyme shape change

• Isoenzymes
– Structurally different enzymes that catalyze the same reaction
• Multi molecular form
• Similar catalytic activity
• Differing biochemical or immunological characteristics
• Can detect by different electrophoresis patterns, absorption patterns, or reaction with specific antibodies

• Cofactor
– Non-protein substances required for normal enzyme activity
– Types
• Activator: inorganic material such as minerals
– (Ca 2+ , Fe 2+)
• Co-enzymes: organic in nature
– (ATP, ADP, nicotinamide)

• Reactions occur spontaneously if energy is available
• Enzymes lower the activation energy for the chemical reactions

• Activation energy
– Excess energy that raises all molecules at a certain temperature to the activation state

• Basic reaction
– S + E ES E + P
– Where
• S= substrate
– Substance on which the enzyme acts
• E= Enzyme
• ES= enzyme-substrate product
– Physical binding of a substrate to the active site of enzyme
• P= Product

• Enzymes differ in their ability to react with different substrates
– Absolute specificity
• Enzyme combines with only one substrate and catalyzes one reaction
– Group specificity
• Combine with all substrates containing a specific chemical group
– Bond specificity
• Enzymes specific to certain chemical bonds
– Stereoisomerism
• Enzymes that mainly combine with only one isomer of a particular compound

• Relationship of the reaction velocity/rate to the substrate concentration
• The Michealis-Menten
Constant (Km)
• The substrate concentration in moles per liter when the initial velocity is ½ V max.
Michaelis-Menten Curve

• First order kinetics
– Rate is directly proportional to substrate concentration
• Zero order kinetics
– Plateau is reached
– depends only on enzyme concentration

• Equation used to distinguish different kinds of inhibition
• Where
– V
0
: velocity/rate of enzymatic activity
– V max
: The maximal rate of reaction when the enzyme is saturated
– K m
: (constant)the substrate concentration that produces ½ of the maximal velocity
– S: substrate concentration

• Adaptation of
Michaelis-Menten equation
• Yields a straight line

Influencing Factors on Enzymatic
Reactions
• Substrate Concentration
• Enzyme Concentration
– The higher the enzyme level, the faster the reaction
• pH
– Most reaction occur in range of 7.0-8.0
– Changes in pH can denature an enzyme
• Temperature
– Most reactions performed at 37 o C
– Increasing temp increases rate of reaction
– Avoid high/low temps due to denaturation of enzyme
• Cofactors
– Influence the rate of reaction
• Inhibitors
– Presence can interfere with a reaction can be reversible or irreversible

• Competitive
– Any substance that competes with the substrate for the active binding sites on the substrate
– Reversible
• Non-competitive
– Any substance that binds to an allosteric site
• Uncompetitive
– Inhibitors bind to the ES complex
– No product produced

Noncompetitive
Inhibition
Irreversible
Inhibition
Competitive
Inhibition

Competitive
Noncompetitive
Uncompetitve

• Historical
– ID of individual enzymes was made using the name of the substrate that the enzyme acted upon and adding “ase” as the suffix
– Modifications were often made to clarify the reaction
– International Union of Biochemistry (IUB) in 1955 appointed a commission to study and make recommendations on nomenclature for standardization

• Components
– Systematic name
• Describes the nature of the reaction catalyzed
• Example: alpha 1,4-glucagon-4-gluconohydrolase
– Recommended name
• Working or practical name
• Example: amylase
– Numerical code
• First digit places enzyme in a class
• Second and third digit represent subclass(s) of the enzyme
• Fourth digit specific serial number in a subclass
• Example: 3.2.1.1

• Standard Abbreviated name
– Accompanies recommended name
– Example: AMS
• Common Abbreviated name
– Example: AMY

• Plasma vs. non-plasma specific enzymes
– Plasma specific enzymes have a very definite/ specific function in the plasma
• Plasma is the normal site of action
• Concentration in plasma is greater than in most tissues
• Often liver synthesized
• Examples: plasmin, thrombin

– Non-plasma specific enzymes have no known physiological function in the plasma
• Some are secreted in the plasma
• Increased number of this type seen with cell disruption or death

• Six classes
– Oxidoreductases
• Involved in oxidation-reduction reactions
• Examples: LDH, G6PD
– Transferases
• Transfer functional groups from one substrate to another
• Examples: AST, ALT
– Hydrolases
• Catalyze the hydrolysis of various bonds
• Examples: acid phophatase, lipase

– Lyases
• Catalyze removal of groups from substrates without hydrolysis, product has double bonds
• Examples: aldolase, decarboxylase
– Isomerases
• Involved in molecular rearrangements
• Examples: glucose phosphate isomerase
– Ligases
• Catabolism reactions with cleavage of ATP
• Example: GSH

• Bishop, M., Fody, E., & Schoeff, l. (2010). Clinical Chemistry:
Techniques, principles, Correlations. Baltimore: Wolters
Kluwer Lippincott Williams & Wilkins.
• http://regentsprep.org/Regents/biology/units/homeostasis/p rocesses.cfm
• http://student.ccbcmd.edu/~gkaiser/biotutorials/proteins/fg9
.html
• Sunheimer, R., & Graves, L. (2010). Clinical Laboratory
Chemistry. Upper Saddle River: Pearson .