FIII: Chymsotrypsin: (Slides 1 - 20) Flashcards
What are ways that enzymes can increase reaction rates?
- Increasing the proximity - holding the reactants close together (Z increases) and the orientation (p)
- Using Chemical Catalysts
Nucleophile Catalysts (lose electrons)
Electrophile Catalysts (gain electrons)
Acid Catalysts (Donate a proton)
Base Catalysts (Gain a proton) - Keep the transition state stable (lower activation energy)
What is Chymotrypsin?
Chymotrypsin is a protease (enzyme) that has the ability to hydrolyze the peptide bonds of the aromatic amino acids: Phe, Tyr, Trp.
Where does Chymotrypsin ‘weakly’ bind?
Chymotrypsin binds ‘weakly’ due to hydrogen bonds to the peptide chain (which acts as the substrate), upstream of the target amino acid
What Terminus does chymotrypsin bind to?
Chymotrypsin will bind to the C-terminus (carboxylate group) of the Aromatic Amino Acids: Phenylalanine. Tyrosine and Tryptophan.
When will Chymotrypsin not bind to these aromatic amino acids?
If any of these amino acids are followed by a proline.
Where does strong binding of chymotrypsin occur?
Strong binding occurs in the binding pocket, when the benzene ring of these amino acids fit into the hydrophobic pocket, so the substrate (peptide chain) binds more tightly.
What is the polarity of chymotrypsins binding pocket?
It is hydrophobic (non-polar)
What does the binding pocket increase?
Proximity, increases Z as it binds the peptide chain more closely together (makes a good fit)
What happens after the substrate (peptide chain) makes a good fit?
The peptide bond immediately following the target amino acid is going to be positioned right next to the catalytic unit
What does the peptide chain following the target amino acid being placed right next to catalytic unit increase?
The orientation (increases P)
What does H2O do?
H2O acts as a nucleophile (wants to give electrons) and attacks the electron deficient carbon, to donate its electrons.
What happens to the Oxygen from the H20 that donated its electrons?
It becomes neutral (the oxygen is not a good nucleophile), making a unfavourable O+ transition state (becomes positive) and forming a bond with the C
What does the Carbon that gets attacked by the nucleophile do?
It remains its stable 8 valence electrons, by giving the electrons to the upper O and withdrawing one of the bonds (no longer double bonded to upper O)
How does the structure of the central carbon change?
It goes from sp2 (trigonal planar) to sp3 (tetrahedral)
What is the transition state?
A semi-stable oxyanion (O negative upper bound C) and the carbon going from sp2 to sp3.
How does the transition state breakdown?
The oxyanion (negative O) returns the bond to the carbon (reforms the double bond) and the carbon sends the electrons to the Nitrogen which forms the leaving group
What happens to the Nitrogen leaving group?
It gains a proton from the surrounding environment and becomes NH2
What is the second pathway that the transition state breakdown can take?
The oxyanion can return its electrons to the carbon, which then can return them to the carbon, then back to the original oxygen from the water causing the water molecule to break off (act as a leaving group) and no reaction occurs.
What does peptide hydrolysis with H20 without a catalyst mean?
The enzyme chymotrypsin is not helping
Why is hydrolysis by neural H2O not good?
- The H20 is not a good nucleophile as the oxygen is too electronegative
- The H20 makes a good leaving group, which can cause no reaction to occur.
How slow is the hydrolysis with neutral H2O?
About one reaction every 10 years (excessively slow)
What are the two easy steps for how Chymotrypsin can speed up the hydrolysis pf a peptide bond?
- Chymotrypsin uses a nucleophilic group in the enzyme to attack the C = O bond.
- Breaks the peptide bond releasing the C-terminal half of the group.
- The N-terminal half remains covalently bonded to the enzyme group (acyl-enzyme intermediate) - Brings in H2O to release the N-terminal and restore the enzyme group to its original state.
How many reactions per second can happen using chymotrypsin?
40 reactions per second.
What is a Catalytic triad?
The catalytic triad is the three amino acids that line up side by side to produce a better nucleophile.
