Lecture 11: enzyme mechanisms Flashcards
TPCK is identified in the active site of _____ and helped to identify what
chymotrypsin; His
catalytic triad
- Ser-195
- His-57
- Asp-102
role of oxygen of Ser-195
oxygen acts as an essential nucleophile because of the pull on its proton from His
reactivity of reaction indirectly caused by _____
negative charge on Asp
the 1st tetrahedral intermediate is stabilized by ____ which stabilizes the t-state (higher/ lower) energy, causing the reaction rate to (increase/ decrease) [the ∆G‡ (increases/ decreases)]
H-bonding interactions with the oxyanion hole; lower energy; reaction rate increases; ∆G‡ decreases
what causes cleavage of the C-N bond? what happens to the N-terminal of the peptide?
the donation of a proton from His [acid] to the nitrogen; creates an acyl bond between N-terminal peptide and serine
how does the 2nd peptide fragment get released from the enzyme? (4)
- His acts as a base and takes a proton from a water molecule
- water attacks the acyl ester carbonyl O
- tetrahedral intermediate formed
- His acts as acid and donates proton to Ser = breaks bond between N-terminal and Ser
the 2nd tetrahedral intermediate is stabilized by ____ with the oxyanion hole
enthalpic interactions
why does steady state take longer than burst phase?
steady state takes longer = dissociation of C-O bond [between Ser and N-terminus] takes longer to reset the enzyme
serine proteases like chymotrypsin (3)
- trypsin
- elastase
- AChE
divergent evolution
evolved from the same ancestral protease
unlike chymotrypsin
subtilisns
convergent evolution
evolved separately to same mechanism
substrate specificity of chymotrypsin
bulky R group, hydrophobic (phenylalanine, tryptophan, tyrosine)
substrate specificity of trypsin
positively charged R group (lysine, argenine)
