Transcript Lecture 5 - enzyme reactions.ppt

Energy Landscape
Product
Reactant
EcoRV
Phosphodiester hydrolysis
H2O
“Reactant State” Crystal Structure
Ala 92
(Mutated
from Lys)
Cleavage
Site
3.7Å
Water
Oxygen
PETRA
IMHOF
Molecular Dynamics of Wild Type
Lys 92
Water
Molecule
Molecular Dynamics of Wild Type
MD I
PETRA
IMHOF
MD II
Principal Component
Principal Components I
Molecular Dynamics of Wild-Type
Deprotonated
Lys 92
Water
Molecule
Muscle Contraction
Thin filament
Thick filament
SONJA SCHWARZL
ATP Hydrolysis by Myosin
Serine Protease
Melanie Böttcher
Index
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Definition
Digestive enzymes
Blood clotting
Complement system
Serine Protease
• Cutting of certain peptide
bonds in other proteins
• Activity depends on a set
of amino acid residues in
the active site of the
enzyme
• Based on nucleophilic
attack of the targeted
peptidic bond by a serine
Serine
• Amino acid
• alcohol as residue
Digestive enzymes
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Chymotrypsin, Trypsin, Elastase
closely-similar structures
different substrate specificities
Pancreatic proteases
 proenzymes trypsinogen and
chymotrypsinogen
 synthesized in the pancreas and
secreted into the lumen of the
small intestine
Mechanism
Active site
Trypsinogen-Trypsin
• loss of six amino acids from one end
• overall structures remain similar
• Ca++ for thermal stability
activated
enzyme does
have more of its
structure
organized into
sheets
Trypsin
• Trypsin cleaves peptide bonds on
the C-terminal side of arginines
and lysines
 Alcaline amino acids
• Activation by enteropeptidase or
through protoelyse
• Optimum pH 7 to 8
Chymotrypsinogen-Chymotrypsin
• Chain is clipped in four places
 release of 2 dipeptides
 creation of two breaks in the backbone
• 1-13 segment retained as part of the active
enzyme, linked on by a disulfide bond
Activation by
trypsin
Chymotrypsin
• Active site: His57-Ser195-Asp102
• Chymotrypsin cuts on the Cterminal side of tyrosine,
phenylalanine, and tryptophan
residues
• hydrolysis of bonds of leucyl,
methionyl, asparaginyl and
glutamyl residues.
Elastase
• cuts peptide bonds next to small, uncharged
side chains such as those of alanine, serine,
valine and threonin
• cuts collagen
Blood Clotting System
• Cascade of several mechanism
Involved Serine Proteases
Thrombin
Plasmin
Factor 10 a
Factor 11 b
Thrombin
• Inactivated Prothrombin
cleaves into activated
Thrombin
• Thrombins cuts
Paraglobuline (=
fibrinoplastic substance)
and forms the soluble
state of it
• Precipitation by
plasmasalt in unsoluble
state
Human Thrombin with the Amino acids
55-65 of Hirudin
Serpins
• Serine Protease Inhibitors
• inhibit the action of their respective serine protease
 serine protease binds the serpin instead of its
normal substrate
 protease makes a cut in the serpin leading to
– the formation of a covalent bond linking the two molecules
– a massive allosteric change in the tertiary structure of the
serpin which moves the attached protease to a site where it
can be destroyed
Serine Protease
Serpin
Chymotrypsin
alpha-1-antichymotrypsin
Complement factor C1s
C1 Inhibitor (C1INH)
Elastase (secreted by neutrophils)
alpha-1-antitrypsin
Clotting factor 10 (X)
Thrombin
antithrombin III
antithrombin III
Plasmin
alpha-2-antiplasmin
Trypsin
pancreatic trypsin inhibitor
Occurrence/ Function
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stop proteolytic activity
blood plasma
clotting
complement
systems where a tiny initial activating event
leads to a rapidly amplifying cascade of
activity
Serpin Deficiencies
• inherited diseases
• mutation in the encoding gene for serpin
• examples
Alpha-1-Antitrypsin
• Serpin for Elastase
• secreted by neutrophils (lung
infection)
emphysema
liver damage
• tests are running
The Others
• Other Serine
Proteases
– Subtilisin
– Acetylcholinesterase
• Serpinlike Molecules
– Angiotensinogen
– Chicken Ovalbumin