Catalytic Strategies Flashcards
EC1 - Oxiodoreductase
Oxidation/Reduction reaction catalyst - always will require an electron carrier such as NAD+
EC2 - Transferases
Transfer a functional group - commonly uses methyl or phosphate
EC3 - Hydrolases
hydrolysis of bonds - uses water to break bonds - all proteases
EC4 - Lysases
Non-hydrolytic non-oxidative breaking of bonds - easiest to think of as splitting a molecule into two pieces
EC5 - Isomerases
Catalyze isomerization changes w/in a single molecule - rearrange the inside of one molecule to a different structure
EC6 - Ligases
Join two molecules by covalent bonds - example if argininosuccinate synthetase
What are enzymes for?
Speed up reactions
Proteases
Enzymes that break peptide bones
Serine Protease contain what?
A residue essential for catalysis
Serine Proteases do what
Cleave peptide bonds at specific sites for cutting. They recognize specific amino acids and cut adjacent to them.
What is the burst phase?
It is when a peptide becomes broken and we see a color reaction
What happens during the long phase?
That is when the other part of the polypeptide chain becomes attached to the enzyme to carry out the remainder of the reaction.
catalytic triad
Three parts- in serine proteases - serine, histidine and aspartic acid. important to remember as it is folded together to create the active site.
What are the steps of the serine protease mechanism - Part 1
Binding of substrate to the S1 pocket - leads to structural change and brings the triad closer together to start reaction
S1 Pocket
region of enzyme that determine what substrate the enzymes binds to
What are the steps of the serine protease mechanism - Part 2
Formation of alkyoxide ion - Histidine moves away from aspartic acid due to electron sink (lots of electrons that are same charge) and pushes towards the serine. This then pulls the proton on serine away and towards the N-group on the Histidine. Once Proton is on N of histidine, this is the alkyoxide
What are the steps of the serine protease mechanism - Part 3
Nucleophilic attack occurs on the carbonyl carbon of the peptide bond, allowing for the oxyanion hole to provide stability to the unstable intermediate.
oxyanion hole
provides environment for unstable intermediate to fall apart. Stabilizes unstable intermediate
What are the steps of the serine protease mechanism - Part 4/5
Breakage and release of the peptide bond and half of the polypeptide chain falls off. This signifies the burst phase on the graph. Other half is covalently linked to the Serine
What are the steps of the serine protease mechanism - Part 6
Water then enters the active site and Nitrogen will attack the proton of water, and Hydrogen will attack the carbonyl carbon. This will create another unstable intermediate which oxyanion hole will stabilize.
The Oxygen attracts the H on the N-ground, breaking the bond on the serine
What are the steps of the serine protease mechanism - Part 7
Release of peptide 2 from the enzymes - end of the long phase
S1 Pocket specificity
Nature of the S1 pocket will determine specificity of the enzyme
Chymotrypsin
Broad, nonpolar - likes phenylalanine, tryptophan and tyrosine
Trypsin
Negatively charged bottom - likes lysin and arginine
Elastase
Narrow, nonpolar - likes glycine, alanine and valine
If we alter the site we see what?
We see that Serine and Histidine are the most important pieces to the reaction. We also see that with no enzyme, there is no feasible reaction time.
Cleavage Agents - Subtilisin
Cuts at C-Terminal of large uncharged side chains
Cleavage Agents - Chemotrypsin
Cuts at C-Terminal side of aromatics (Phe,Tyr,Trp)
Cleavage Agents - Trypsin
Cuts at C-Terminal side of Lysin and Arginines
Cleavage Agents - Carboxypeptidase
Cuts at N-Terminal of C-terminal amino acid
Cleavage Agents - Elastase
hydrolyzes C-side of small AA’s (Gly, Ala)
Cleavage Agents - Cyanogen Bromide
Chemical - Hydrolyzes C-side of Met
Serpins
Serine Protease Inhibitors - polypeptides that block access to active site to prevent other binding from occurring - important for medicine
alpha-1-antitrypsin
inhibits elastase - made by body to protect lungs - w/o this this leads to emphysema.
Reactive Oxygen Species
Oxygen byproduct
alkyoxide ion
This is very reactive and responsible for the reaction that occurred
