Mechanism of action
MUDr. Martin Vejražka, PhD.
Can organic reactions run outside the cell?
Justus von Liebig http://de.wikipedia.org/wiki/Justus_von_Liebig
Louis Pasteur http://it.wikipedia.org/wiki/Louis_Pasteur
Hans Büchner 1897
• Filtered yeast juice can metabolise sugar
• ἐν ζυµῇ
= in yeast
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James Sumner 1926
• Enzymes are proteins http://en.wikipedia.org/wiki/James_B._Sumner
• Ribozymes – catalytically active
RNA
Enzymes
• Small amount
• Not changed irreversibly in reaction
– i.e. can be recycled
• Do not change thermodynamics of reaction
– K, ∆ G
• Increase the reaction rate
Catalytic activity
Without catalyst
(per year)
„Classical“ catalyst
(per year)
100-1000×
Enzyme
(per second)
10 8 -10 23 ×
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Stability and reactivity
≠
Decreased activation energy
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Intermediary state
Bacterial prolinracemase:
L-Pro Intermediary state D-Pro
• Compounds similar to intermediary state frequently act as inhibitors
Enzyme-substrate complex
• Active site
• Non-covalent interactions
– Ionic
– Hydrogen bridges
– Hydrophobic
Entry of the substrate to the active center
• Dehydration of the substrate
• Binding of the substrate to the surface of the enzyme
• Reaction of the substrate
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Binding the substrate to the enzyme
• Lock and key
• Induced fit
Catalysis by proximity
• High local concentration
• Orientation of the substrates
• Low entropy
Acid-base catalysis
• High concentration of acidic or basic groups in the active center
– Aminoacids of the enzyme
– Prosthetic groups
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Catalysis by tension
• Covalent bond under tension, more susceptible to break-down
• Stabilisation of intermediary state
• „Lytic“ reactions
Covalent catalysis
• A functional group covalently bond to the enzyme
• E.g. transaminases
• Often „ping-pong“ mechanism
• In active center:
– Cys
– Ser
– (His)
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Enzyme specifity
• Substrate
– Strict for optic isomers
– Less strict to length of chain
• Reaction
Composition of an enzyme
• Holoenzyme
– Apoenzyme
– Non-proteinic part
• Prosthetic group
• Co-factor
• Co-enzyme
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Prosthetic group
• Covalently bond to the apoenzyme
• Metals
– About 1/3 of enzymes (metaloproteins)
– Often in
• Hem
• FeS-clusters
Prosthetic group
• Binding and orientation of the substrate
• Covalent binding of intermediates
– E.g. Co 2+ in vit. B
12
• Electrophilic / nucleophilic
Co-factors
• Can be dissociated
• Must be present near the enzyme
• Metalic ions
– Metal activated enzymes
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Co-enzymes
• More complex organic compounds
– Often derived from vitamins
• Serve as carriers for functional groups and substrates
• Stabilise high-reactive groups
(e.g. H)
Name of an enzyme
• -ase
• International Union of Biochemistry
– Name and code
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Names of enzymes
• Hexokinase
• ATP:
D
-hexose-6-phosphotransferase
• E.C. 2.7.1.1
– 2 = transferases
– 7 = transfer of phosphate
– 1 = alcohol is the acceptor
– 1 = serial number
Isoenzymes
• Different enzymes catalysing the same reaction
• Different localisation
• Different regulation
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Enzyme activity
• Katal, kat
– The amount of enzyme that catalyses conversion of 1 mol of the substrate per second
• International unit, U or IU
– 1 mmol per 1 min
Enzyme activity
• Specific activity
– kat / mg prot.
• Activity concentration
– kat / L
Mechanism of action
MUDr. Martin Vejražka, PhD.
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