Download Enzymes - Dr. Hamad Ali Yaseen

11/18/2014
Enzymes
Hamad Yaseen, PhD
Biochemistry 210
[email protected]
MLS Dept, FAHS, HSC, KU
Read Chapter 23
Enzymes as Biological Catalysts
• Enzymes are proteins
that increase the rate
of reaction by
lowering the energy of
activation. Without
themselves
undergoing any
changes.
• They catalyze nearly
all the chemical
reactions taking place
in the cells of the body
• Enzymes have unique
three-dimensional
shapes that fit the
shapes of reactants
(substrates)
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Rate Enhancement
Formation of carbonic acid from CO2 and H2O.
Uncatalyzed rate = 1.3 x 10-1 molecules/sec
Catalyzed rate = 1.0 x 106 molecules/sec
Rate enhancement = 7.7 x 106 times
Carbonic
Anhydrase
Table of Rate Enhancements
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Enzymes
• The vast majority of enzymes are proteins.
• However, proteins are not the only biological catalysts.
• Ribozymes, are enzymes made of ribonucleic acid.
Naming Enzymes
• The name of an enzyme identifies the reacting substance
- usually ends in –ase
• For example, sucrase catalyzes the hydrolysis of sucrose
• The name also describes the function of the enzyme
• For example, oxidases catalyze oxidation reactions
• Sometimes common names are used, particularly for the
digestion enzymes such as pepsin and trypsin
• Some names describe both the substrate and the function
• For example, alcohol dehydrogenase oxides ethanol
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Enzyme Nomenclature
• active site - a region of an enzyme comprised of different amino acids where
catalysis occurs (determined by the tertiary and quaternary structure of each
enzyme)
• substrate - the molecule being utilized and/or modified by a particular enzyme at
its active site
• co-factor - organic or inorganic molecules that are required by some enzymes for
activity. These include Mg2+, Fe2+, Zn2+ and larger molecules termed co-enzymes
like nicotinamide adenine dinucleotide (NAD+), coenzyme A, and many vitamins.
• prosthetic group - a metal or other co-enzyme covalently bound to an enzyme
• holoenzyme - a complete, catalytically active enzyme including all co-factors
• apoenzyme - the protein portion of a holoenzyme minus the co-factors
• isozyme - (or iso-enzyme) an enzyme that performs the same or similar function
of another enzyme. This generally arises due to similar but different genes
encoding these enzymes and frequently is tissue-type specific or dependent on
the growth or developmental status of an organism.
Enzyme structure
• Enzymes are proteins
• They have a globular shape
• A complex 3-D structure
• Human pancreatic amylase
© 2007 Paul Billiet ODWS
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The active site
• One part of an enzyme, the
active site, is particularly
important
• The shape and the chemical
environment inside the active site
permits a chemical reaction to
proceed more easily
© H.PELLETIER, M.R.SAWAYA
ProNuC Database
© 2007 Paul Billiet ODWS
Cofactors
• An additional non-protein
molecule that is needed by some
enzymes to help the reaction
• Tightly bound cofactors are
called prosthetic groups
• Cofactors that are bound and
released easily are called
coenzymes
• Many vitamins are coenzymes
Nitrogenase enzyme with Fe, Mo and ADP cofactors
Jmol from a RCSB PDB file © 2007 Steve Cook
H.SCHINDELIN, C.KISKER, J.L.SCHLESSMAN, J.B.HOWARD, D.C.REES
STRUCTURE OF ADP X ALF4(-)-STABILIZED NITROGENASE COMPLEX AND ITS
© 2007 Paul Billiet ODWS
IMPLICATIONS FOR SIGNAL TRANSDUCTION; NATURE 387:370 (1997)
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Classification of Enzymes
• Enzymes are classified according to the type of reaction they
catalyze:
Class
 Oxidoreductases
 Transferases
 Hydrolases
 Lyases
 Isomerases
 Ligases
Reactions catalyzed
Oxidation-reduction
Transfer groups of atoms
Hydrolysis
Add atoms/remove atoms to/from a double bond
Rearrange atoms
Use ATP to combine molecules
Oxidoreductases, Transferases and Hydrolases
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Lyases, Isomerases and Ligases
Active Site of an Enzyme
• The active site is a region
within an enzyme that fits
the shape of substrate
molecules
• Amino acid side-chains align
to bind the substrate
through H-bonding, saltbridges, hydrophobic
interactions, etc.
• Products are released when
the reaction is complete
(they no longer fit well in the
active site)
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Enzyme Specificity
• Enzymes have varying degrees of specificity for substrates
• Enzymes may recognize and catalyze:
- a single substrate
- a group of similar substrates
- a particular type of bond
Factors influence enzyme activity?
• Enzyme and substrate concentration
• Temperature
• P.H
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Enzyme Concentration and Reaction Rate
• The rate of reaction increases as enzyme concentration
increases (at constant substrate concentration)
• At higher enzyme concentrations, more enzymes are available
to catalyze the reaction (more reactions at once)
• There is a linear relationship between reaction rate and
enzyme concentration (at constant substrate concentration)
Substrate Concentration and Reaction Rate
• The rate of reaction increases as substrate concentration
increases (at constant enzyme concentration)
• Maximum activity occurs when the enzyme is saturated
(when all enzymes are binding substrate)
• The relationship between reaction rate and substrate
concentration is exponential, and asymptotes (levels off)
when the enzyme is saturated
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Temperature and Enzyme Activity
• Enzymes are most active at an optimum temperature
(usually 37°C in humans)
• They show little activity at low temperatures
• Activity is lost at high temperatures as denaturation occurs
pH and Enzyme Activity
• Enzymes are most active at optimum pH
• Amino acids with acidic or basic side-chains have the proper
charges when the pH is optimum
• Activity is lost at low or high pH as tertiary structure is
disrupted
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Optimum pH for Selected Enzymes
• Most enzymes of the body have an optimum pH of about 7.4
• However, in certain organs, enzymes operate at lower and
higher optimum pH values
What are the mechanisms of enzyme action?
• Lock-and-key model
• Induced fit model
• Catalytic power of enzymes
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Lock-and-Key Model
• In the lock-and-key model of enzyme action:
- the active site has a rigid shape
- only substrates with the matching shape can fit
- the substrate is a key that fits the lock of the active site
• This is an older model, however, and does not work for all
enzymes
Lock & Key – Fischer (1894)
A proposal for ES
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Induced Fit Model
• In the induced-fit model of enzyme action:
- the active site is flexible, not rigid
- the shapes of the enzyme, active site, and substrate adjust to
maximumize the fit, which improves catalysis
- there is a greater range of substrate specificity
• This model is more consistent with a wider range of enzymes
Induced Fit – Koshland (1963)
A proposal for ES
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Catalytic power of enzymes
• Previous models emphasized on the shape of the active site.
However the chemistry at the active site is actually the most
important factor.
Enzyme Catalyzed Reactions
• When a substrate (S) fits properly in an active site, an
enzyme-substrate (ES) complex is formed:
E + S  ES
• Within the active site of the ES complex, the reaction occurs
to convert substrate to product (P):
ES  E + P
• The products are then released, allowing another substrate
molecule to bind the enzyme
- this cycle can be repeated millions (or even more) times
per minute
• The overall reaction for the conversion of substrate to
product can be written as follows:
E + S  ES  E + P
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Example of an Enzyme Catalyzed Reaction
• The reaction for the sucrase catalyzed hydrolysis of sucrose to
glucose and fructose can be written as follows:
E + S  ES  E + P1 + P2
where E = sucrase, S = sucrose, P1 = glucose and P2 = fructose
Isoenzymes
• Isoenzymes are different forms of an enzyme that catalyze
the same reaction in different tissues in the body
- they have slight variations in the amino acid sequences of
the subunits of their quaternary structure
• For example, lactate dehydrogenase (LDH), which converts
lactate to pyruvate, consists of five isoenzymes
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How are Enzymes Regulated?
• Feedback Control:
• An enzyme regulation process in which formation of a product inhibits an
earlier reaction in the sequence.
• The reaction product of one enzyme may control the activity of another,
especially in a complex system in which enzymes work cooperatively.
How are Enzymes Regulated?
• Proenzymes
• Some enzymes are manufactured in the body in an inactive form. To make
them active, a small part of their polypeptide chain must be removed.
• Why this is important? Think trypsin
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How are Enzymes Regulated?
• Allosterism
• An event that occurs at a site other than the active site but that eventually
affects the active site.
• Any enzyme regulated by this mechanism is called Allosteric enzyme.
• These could be: Negative modulators and positive modulators
Enzyme Inhibitors
• Inhibitors (I) are molecules that cause a loss of
enzyme activity
• They prevent substrates from fitting into the active
site of the enzyme:
E + S  ES  E + P
E + I  EI  no P formed
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Reversible Inhibitors (Competitive Inhibition)
• A reversible inhibitor goes
on and off, allowing the
enzyme to regain activity
when the inhibitor leaves
• A competitive inhibitor is
reversible and has a
structure like the substrate
- it competes with the
substrate for the active site
- its effect is reversed by
increasing substrate
concentration
Reversible Inhibitors (Noncompetitive Inhibition)
• A noncompetitive inhibitor has
a structure that is different
than that of the substrate
- it binds to an allosteric site
rather than to the active site
- it distorts the shape of the
enzyme, which alters the shape
of the active site and prevents
the binding of the substrate
• The effect can not be reversed
by adding more substrate
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How are Enzymes Regulated?
• Protein Modification
• The modification is usually change in primary structure, typically by addition
of a functional group covalently bound to the apoenzyme.
• A well described example: Phosphorylation
How are Enzymes Regulated?
• Isoenzymes
• Regulatory of enzyme activity via having more than one form of the enzymes
in different tissues and organs.
A good example is Lactate dehydrogenase (LDH).
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Isoenzymes
• Why this is important? “The heart example”
• The heart is purely aerobic organ.
• LDH is used to convert lactate into pyruvate.
• The H4 enzyme is allosterically inhibited by high levels of pyruvate (its
product).
• Has a higher affinity for lactate (its substrate) than M4 enzyme, which is
optimized for the opposite reaction. The M4 isoenzyme favors the
production of lactate.
Diagnostic Enzymes
• The levels of diagnostic enzymes in the blood can be used
to determine the amount of damage in specific tissues
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