chymotrypsin poster

References: Kashima A, Inoue Y, Sugio S, Madea I, Nose T and Shimohigashi Y (1998): X-ray crystal structure of a dipeptide-chymotrypsin complex in an inhibitory interaction; Khan A and James N.G. (1998): Molecular mechanisms for the conversion of zymogens to active proteolytic enzymes; Latha B, Ramakrishnan M, Jayaraman V, Babu M (1997): Serum enzymatic changes modulated using trypsin: chymotrypsin preparation during burn wounds in humans. Burns, 23:560-4; Polgar L (2005): The catalytic triad of serine proteases; Wenzhe Ma, Chao Tang, and Luhua Lai (2005): Specificity of Trypsin and Chymotrypsin: Loop-Motion-Controlled Dynamic Correlation as a Determinant. Biophysical Journal. Volume 89. 1183–1193.

2. Structure •  Three chains linked by disulfide bonds (Refer Fig. 3) •  Three factors which make construct the active site of the chymotrypsin

1. Catalytic triad: There is concerted hydrogen bonding between the residues of the triad (Polgar, 2005): the side chain of Ser-195 is hydrogen bonded to the imidazole of the His-57, whilst the the –NH group of this imidazole is hydrogen bonded to the carboxylate group of Asp-102 (Refer Fig. 1)

Ø  His-57 acts as a general base to increase nucleophilicity of the O atom in Ser-195 (Refer to Section 4, Step 2)

2. S1 primary pocket: Only substrates with aromatic residues

can bind here (Refer Fig. 3) gives chymotrypsin its primary specificity

3. Oxyanion hole: Amide nitrogen from peptide backbone of Ser-195 and Gly-193 help stabilise:

Ø  Unstable tetrahedral intermediate (Refer Step 3 in the

mechanism)

Ø  Transition state that proceeds formation to tetrahedral

intermediate (Wenzhe et al, 2005; Polgar, 2005)

1. Introduction

•  Chymotrypsin belongs to a superfamily of serine proteases involved

in hydrolysis of peptide bonds using an active serine residue that is

part of a “catalytic triad”: Asp-102, His-57, Ser-195

•  Located in the pancreas - vital for the digestion of dietary proteins

•  It has a primary specificity for large, aromatic, hydrophopbic amino

acid residues (Phe, Tyr, Trp) (Wenzhe et al, 2005)

3. Regulation of Chymotrypsin

•  Due to the power of proteolytic activity, premature hydrolysis must be

avoided. Chymotrypsin is initially synthesised as a zymogen called

chymotrypsinogen (Khan et al, 1998). This zymogen is activated by

proteolytic cleavage the overall structure (Refer Fig. A)

4. Catalytic Mechanism of Chymotrypsin STEP I – ACYLATION (Polgar, 2005)

1.  Substrate positioned within the active site 2.  O atom on Ser-195 (Fig. 1) induces a nucleophilic

attack of Ser-195 to the carbon atom within the carbonyl of the peptide bond

3.  Unstable tetrahedral intermediate formed Ø  The transition state converts to a high energy

tetrahedral intermediate 4.  An acyl-enzyme is formed as the His-57 acts as a

general base 5.  An amine compound is the leaving molecule group

due to the peptide cleavage that occurs in Step 4 ------------------------------------------------------------------------

STEP II – DEACYLATION (Polgar, 2005)

6.  Water molecule binds onto the active site 7.  His-57 now acts a general acid by drawing a proton away

from a water molecule 8.  Ester group in acyl enzyme is hydrolysed 9.  The O atom in H2O is a strong nucleophile 10.  Repeat Step 3-4 11.  A carboxylic acid compound leaves and the enzyme is

ready for the next set of catalysis

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5. Future Research •  Treatment of burns by the decreasing tissue destruction •  Treatment of hand fractures to reduce redness and

inflammation (Latha et al, 1997)

Fig. 2 The overall structure of chymotrypsin emphasising The S1 pocket is located near the catalytic triad and Gly-193 (in green) which is part of the oxyanion hole. PDUB 7GCH.

Fig.1 Catalytic Triad: Ser-195, His-57 and Asp-102. The dashed lines demonstrate the hydrogen bonding between the residues of the catalytic triad . Due to these interactions, the weak nucleophile of O in Ser-195 becomes a stronger nucleophile (Kashima et al, 1998)

S1 specificity pocket (2) Oxyanion hole

Fig. 3 The overall

spherical structure of

chymotrypsin showing

Chains A, B and C. They

are linked by disulfide

bonds, shown in blue

(Kashima et al, 1998;

Khan et al, 1998) Chain A

Chain B

Chain C

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