Studies on the catalytic mechanism and inhibition of serine proteases are w
idely used as paradigms for teaching enzyme catalysis. Ground-breaking work
on the structures of chymotrypsin and subtilisin led to the idea of a cons
erved catalytic triad formed by the active site Ser, His and Asp residues.
An oxyanion hole, consisting of the peptide amide of the active site serine
and a neighbouring glycine, was identified, and hydrogen bonding in the ox
yanion hole was suggested to stabilize the two proposed tetrahedral interme
diates on the catalytic pathway. Here we show electron density changes cons
istent with the formation of a tetrahedral intermediate during the hydrolys
is of an acyl-enzyme complex formed between a natural heptapeptide and elas
tase. No electron density for an enzyme-product complex was observed. The s
tructures also suggest a mechanism for the synchronization of hydrolysis an
d peptide release triggered by the conversion of the sp(2) hybridized carbo
nyl carbon to an sp(3) carbon in the tetrahedral intermediate. This affects
the location of the peptide in the active site deft, triggering the collap
se of a hydrogen bonding network between the peptide and the P-sheet of the
active site.