Conformational changes accompanying the formation of chymotrypsin-substrate complexes. Evidence for the involvement of an N-terminal α-amino group in the activity and the conformation of the enzyme

Implications of X-Ray Crystallographic Studies of Protein Structure

“…The involvement of a histidine (128) and an a-amino group (30)(31)(32) at the active site of a-chymotrypsln also was predicted, q-'itration studies of lysozyme revealed the presence of carboxyl groups with abnormal pK's (129). Crystallographic studies show that the catalytic res:[due glutamlc 35 is very likely one of these carboxyls (11).…”

“…The participation histidine 57 was directly demonstrated by affinity labeling (27,28), as was the proximity of methionine 192 (29). Hess and co-workers (30)(31)(32) emphasized the importance of the charged s-amlno group of isoleucine 16 for the following reasons: (a) The only necessary step in the activation process cleavage of the peptide bond between residues 15 and 16 in chylaaotrypsinogen to yield a new a-amino group on isoleucine 16. (b) The enzyme is catalytically active only if this a-amino group is protonated, An ionizing group 30 STRYER with a pK of about 8.5 controls the formation of productive enzyme-substrate complexes but does not influence the bond-breaking step.…”

“…In chymotrypsin, the only charged residues inside the molecule are an a-amino group and an adjacent aspartate side-chain (14). This ion-pair has been shown to be important in stabilizing the active conformation of the enzyme (30)(31)(32).…”

Ultrasonic studies of proton‐transfer reactions at the catalytic site of α‐chymotrypsin

“…Confirmatory evidence is provided by the fact that at 40°C in a 0.1 M borate buffer of pKa = 9 an ultrasonic absorption peak was observed at pH 8.5, that stems from I1e-16. This peak, indeed, does not exist in the zymogen and disappears in the DFP-treated enzyme, in which the pKa value of Ile-16 is raised to above 10 [6].…”

“…(ii) For Ile-16, because its pKa value is known to be of the order of 8.5 [6], and its absorption peak, therefore, cannot be observed in a buffer of pKa value close to 7. Confirmatory evidence is provided by the fact that at 40°C in a 0.1 M borate buffer of pKa = 9 an ultrasonic absorption peak was observed at pH 8.5, that stems from I1e-16.…”