Enzyme catalytic strategies Flashcards
2 things that enzyme activity depend on?
pH
Temperature
How does increase in temp affect enzyme activity?
Increases it as molecules move faster
4 catalytic principles used by enzymes
Covalent catalysis
General acid-base catalysis
Catalysis by approximation
Metal ion catalysis
Which catalytic strategies does chymotrypsin combine?
Covalent, acid-base and transition-state
What does protease catalyse?
Proteolysis
Cuts down proteins
Difference between nucleophile and electrophile
Nucleophile: Accepts protons and donates electrons
Electrophile: Donates protons and accept electrons
What is chymotrypsin involved in?
Breaking down proteins into small peptides by cutting a specific location on peptide backbone
What is the catalytic triad?
Serine, Histadine and Asperegine
They have to be close in the folding
Where does the cutting occur in chymotrypsin?
In the hydrophobic pocket, the serine cuts the protein when it is attached to the hydrophobic pocket
What does the oxyanion hole do?
Protects the enzyme
Does the S1/hydrophobic pocket differ in protease?
Yes
Do proteases cut in only one point?
Yes
What allows for the specificity in proteases?
The difference in hydrophobic pockets
Example of aspartyl protease
Pepsine
What is site-directed mutagenesis?
Replace one amino acid with another
Example how the catalytic triad was discovered
3 classes of proteases
Cysteine
Aspartyl
Metalloproteases
Which amino acid is conservative replacement (keep good activity) in serine?
Cystine
What is indinavir?
Analogue of the substrate
What does carbonic anhydrase do?
Make a fast reaction faster
Helps with metabolism
How many carbonic anhydrasases (esoenzymes) in the body?
7
What is the Kcat?
Enzyme turn-over number
Amount of product produced per unit of time per enzyme
How does pH effect carbonic anhydrase activity?
Reaction is higher (Kcat) in basic environment because a group that loses a proton plays an important role in the activity of carbonic anhydrase
Which was the first enzyme that uses a metal that was discovered?
Carbonic anhydrase
What is lysozyme?
An antibacterial enzyme
What does lysozyme attack?
Peptidoglycan
What is peptidoglycan?
Polysaccharide found in many bacterial cell walls
Example of where lysozyme I found?
In the numerous body fluid (blood, sweat, saliva, sweat, tears, urine)
What are isoenzymes?
Enzyme variants that are the product of different genes and thus represent
What are allozymes?
different loci
Enzymes that are the product of different alleles of the same gene
Why are isoenzymes clinically relevant?
Helps determine where the enzymes in the tissue are coming from. This helps determine if there are for example liver damage (if there are liver enzymes present in the blood)
How can heart attacks be found out with the help of isoenzymes?
If heart enzymes are found in the blood when isoenzymes are separated by electrophoresis there can be a sign of heart attack
How are different isoenzymes identified?
Electrophoresis (because of charge and size)
What is the same in isoenzymes?
The active site
What kind of enzyme class do proteases belong to?
Hydrolase
Can a nonbiological catalyst produce more than one product with the occurrence of side reactions?
Yes
Must a substrate bind to the active site before catalysis can occur?
Yes
Do enzymes change the free energy of a reaction?
No
What is the optimal temperature for enzyme activity?
35-45c
Is the optimal pH the same for all enzymes?
No it very much depends on the enzyme and its function and where its found
What is covalent catalysis?
Active site contains a nucleophile that is briefly covalently modified
What is general acid-base catalysis?
A molecule other than water donates or accepts a proton
What is catalysis by approximation?
A molecule other than water donates or accepts a proton
What is metal ion catalysis?
Can function in different ways
Example: electrophilic catalyst
What does the hydrophobic pocket recognise?
Peptide bonds
Why does apartyl protease work best in acidic conditions?
Because it can be protenated
Serine, histamine and superfine in the catalytic triad are not close when looking at the primary structure, how do they make up the catlytic triad?
They are close in the tertiary structure thanks to the folding
What in chymotrypsin catalyses the cleavage of peptide bonds ?
The catalytic triad
What explains substrate specificity of trypsin, chymotrypsin and elastase?
Structual difference in the binding site
What is the rocket in chymotrypsin, polar or nonpolar?
Nonpolar
How does the pocket of trypsin differ from that of chymotrypsin?
It has Asp instead of Ser
How does elastase differ from chymotrypsin?
There are no pockets present
Why are there no pockets in elastase?
Because 2 glycerine is replaced by Val and Thr
What does the hydrophobic pocket of chymotrypsin bind?
Aromatic amino acids
Is chymotrypsin hydrophobic or hydrophilic?
Hydrophobic
Why does the group have to be protected to work better in aspartyl proteases?
Because the conditions have to be acidic
What is pepsin?
An aspartyl protease
How do metalloproteases work?
The peptide carbonyl group is attacked by a metal-activated water molecule
What carbonic anhydrase used to do in animals?
Transport CO2 out of muscles and maintain blood pH balance
What catalyses the reaction of hydration CO2 to form bicarbonate?
Carbonic anhydrase
Which ion is found at the active site of carbonic anhydrase?
Zinc ion
What does catalysis with carbonic anhydrase entail?
Zinc activation of a water molecule
What does zinc ion promote the ionisation of? and what does it result in
Bound H2O, resulting nucleophilic OH- attacks carbon of CO2
Why does the zinc in carbonic anhydrase make the reaction much faster?
Because water without zinc is a very weak acid/nucleophile
What is peptidoglycan?
A polysaccharide found in many bacterial cell walls
What does the cleavage of the cell wall of bacteria lead to?
Lysis of bacteria
Antibacterial enzyme
Lysozyme
When is lysozyme released into the blood stream?
When granulocytes and monocytes are released
What does lactate dehydrogenase catalyse?
A step in anaerobic glucose metabolism and glucose synthesis
What does LDH stand for?
Lactate dehydrogenase
Why can isozymes be exerted by electrophoresis?
Because they differ slightly in charge
