Lecture 10 - Acid-base Catalysis

Acid Catalysis

PROTONATE the molecule to make more electrophilic (acid is a proton DONOR)

Base Catalysis

DEPROTONATE water to make more nucleophilic (base is a proton ACCEPTOR)

Amino Acid Bases

Arginine, lysine, histidine, tyrosine, cysteine

Amino Acid Acids

Aspartic acid, glutamic acid, histidine

Importance of histidine in acid-base catalysis

Can act as BOTH acid and base catalyst

What amino acids can participate in acid-base catalysis?

Those with IONIZABLE side chains

Aspartate, glutamate, HISTIDINE, arginine, lysine, cysteine, tyrosine

Graph of acid catalysis

Higher v₀ at lower pH

Graph of base catalysis

Higher v₀ at higher pH

Protease Papain

• Contains cysteine and histidine, which both act in catalysis

  • Cysteine - Base (deprotonated → protonated)

  • Histidine - Acid (protonated → deprotonated)

pH rate profile

Graph that represents ionization states of ALL species in the transition state

Papain unique feature

• Cysteine and histidine are very close together, resulting in change in pKa

  • Histidine wants Cysteine’s proton, trying to pull it

pKa’s change…

Depending on environment (ex. proximity to other residues), altering acid-base behavior

Chymotrypsin

• Endoprotease during digestion

• Cleaves peptide bonds of bulky hydrophobic residues

  • Phe, Try, Tyr, Leu

• Cleaves esters

• SERINE PROTEASE

Is chymotrypsin an endoprotease or exoprotease?

ENDOprotease

What does chymotrypsin cleave?

Esters and bulky, hydrophobic residues

Examples of bulky, hydrophobic residues

Phenylalanine, Tryptophan, Tyrosine, Leucine

Chymotrypsin function

Intestinal digestion

What family does chymotrypsin belong to?

Serine proteases

Key players in chymotrypsin action

1. Serine - reactive nucleophile

2. Aspartic acid - electrostatic interaction, enhances nucleophilicity

3. Histidine - acid/base

4. Oxyanion hole - stabilizes transition

5. Specificity pocket - binds bulky hydrophobic residues

Kinetics of serine protease (chymotrypsin)

Burst and steady-state phases

Burst phase

Beginning of chymotrypsin activity, acylation of chymotrypsin (all enzyme is free)

Steady-state phase

Slow phase, when enzyme is acylated and it takes longer for deacylation to occur

Serine proteases CANNOT function acylated

Size of burst =

Amount of enzyme available

more enzyme = larger burst

Steady-state is proportional to:

Concentration of [E]