Lecture Notes: Serine Proteases and Peptide Bond Hydrolysis

Introduction

• Exploration of a different class of enzymes: Serine Proteases.
• Key feature: An essential serine residue in the active site.
• Color representation:
  • White: Enzyme active site
  • Green: Peptide backbone
  • Red: Water molecules
  • Yellow: Electron flow

Active Site Composition

• Important Amino Acids:
  1. Aspartic Acid
  2. Histidine
  3. Serine
• These amino acids are positioned to interact closely in the enzyme's tertiary structure.

Hydrogen Bonding and Electron Density

• A crucial hydrogen bond exists between aspartic acid and nitrogen on histidine.
• This bond influences electron density, making nitrogen's lone pair more reactive.

Mechanism Overview

Step 1: Nucleophile Creation

• Reactive nitrogen lone pair deprotonates serine.
• Formation of a negatively charged oxyanion on serine, acting as a nucleophile.
• Nucleophile attacks the carbonyl, shifting electrons and forming a tetrahedral intermediate.

Step 2: Proton Shuffling

• Proton shuffling step forms a positively charged nitrogen on peptide backbone.
• Tetrahedral intermediate collapses, breaking the peptide bond.

Step 3: Water Molecule Involvement

• Water replaces the N-terminus; necessary for hydrolysis.
• Water is deprotonated to form a hydroxide anion, a strong nucleophile.
• Hydroxide anion attacks electrophilic carbon, forming another tetrahedral intermediate.

Step 4: Collapsing Tetrahedral Intermediate

• Positive charge creation on oxygen allows the CO bond to break.
• Tetrahedral intermediate collapses, regenerating the active site.

Conclusion

• Complete regeneration of the active site, allowing for subsequent reactions.
• Mechanism includes two complete steps: initiating with serine nucleophile, followed by water.
• Similar themes to previous enzyme-catalyzed hydrolysis mechanisms.
• Key difference: Two-step process with serine and water involvement.

Summary

• Serine proteases facilitate peptide bond hydrolysis through a complex catalytic mechanism involving key amino acids.
• The process regenerates the enzyme's active site, readying it for new substrates.