Chymotrypsin catalytic triad

This endopeptidase [EC 3.4.21.4], a member of the peptidase family SI, hydrolyzes peptide bonds at Arg—Xaa and Lys—Xaa. See Chymotrypsin Catalytic Triad Acyl-Serine Intermediate... 

Lysyl-tRNA phosphatidylglycerol transferase, LYSYLTRANSFERASE a-LYTIC PROTEINASE CHYMOTRYPSIN CATALYTIC TRIAD D-Lyxose,... 

ACYL-SERINE INTERMEDIATE CHYMOTRYPSIN CATALYTIC TRIAD TRYPTOPHANASE TRYPTOPHAN SYNTHASE T state,... 

Scheme 12.66. A cartoon representation of the chymotrypsin catalytic triad in action. Cleavage of a peptide on the carbonyl side of the bond (to phenylalanine [Phe, F], tyrosine [Tyr, Y], or tryptophan) by the triad of Asp (D)-102, His (H)-57, and Ser (S)-195.

Figure 11.7 Schematic diagram of the structure of chymotrypsin, which is folded into two antiparallel p domains. The six p strands of each domain are red, the side chains of the catalytic triad are dark blue, and the disulfide bridges that join the three polypeptide chains are marked in violet. Chain A (green, residues 1-13) is linked to chain B (blue, residues 16-146) by a disulfide bridge between Cys 1 and Cys 122. Chain B is in turn linked to chain C (yellow, residues 149-245) by a disulfide bridge between Cys 136 and Cys 201. Dotted lines indicate residues 14-15 and 147-148 in the inactive precursor, chmotrypsinogen. These residues are excised during the conversion of chymotrypsinogen to the active enzyme chymotrypsin.

Figure 11.9 A diagram of the active site of chymotrypsin with a bound inhibitor, Ac-Pro-Ala-Pro-Tyr-COOH. The diagram illustrates how this inhibitor binds in relation to the catalytic triad, the strbstrate specificity pocket, the oxyanion hole and the nonspecific substrate binding region. The Inhibitor is ted. Hydrogen bonds between Inhibitor and enzyme are striped. (Adapted from M.N.G. James et al., /. Mol. Biol. 144 43-88, 1980.)...

A closer examination of these essential residues, including the catalytic triad, reveals that they are all part of the same two loop regions in the two domains (Figure 11.10). The domains are oriented so that the ends of the two barrels that contain the Greek key crossover connection (described in Chapter 5) between p strands 3 and 4 face each other along the active site. The essential residues in the active site are in these two crossover connections and in the adjacent hairpin loops between p strands 5 and 6. Most of these essential residues are conserved between different members of the chymotrypsin superfamily. They are, of course, surrounded by other parts of the polypeptide chains, which provide minor modifications of the active site, specific for each particular serine proteinase. 

Figure 11.10 Topological diagram of the two domains of chymotrypsin, illustrating that the essential active-site residues are part of the same two loop regions (3-4 and 5-6, red) of the two domains. These residues form the catalytic triad, the oxyanion hole (green), and the substrate binding regions (yellow and blue) including essential residues in the specificity pocket.

The active site of subtilisin is outside the carboxy ends of the central p strands analogous to the position of the binding sites in other a/p proteins as discussed in Chapter 4. Details of this active site are surprisingly similar to those of chymotrypsin, in spite of the completely different folds of the two enzymes (Figures 11.14 and 11.9). A catalytic triad is present that comprises residues Asp 32, His 64 and the reactive Ser 221. The negatively charged oxygen atom of the tetrahedral transition state binds in an oxyanion hole,... 

Figure 11.14 Schematic diagram of the active site of subtilisin. A region (residues 42-45) of a bound polypeptide inhibitor, eglin, is shown in red. The four essential features of the active site— the catalytic triad, the oxyanion hole, the specificity pocket, and the region for nonspecific binding of substrate—are highlighted in yellow. Important hydrogen bonds between enzyme and inhibitor are striped. This figure should be compared to Figure 11.9, which shows the same features for chymotrypsin. (Adapted from W. Bode et al., EMBO /.

Serine proteinases such as chymotrypsin and subtilisin catalyze the cleavage of peptide bonds. Four features essential for catalysis are present in the three-dimensional structures of all serine proteinases a catalytic triad, an oxyanion binding site, a substrate specificity pocket, and a nonspecific binding site for polypeptide substrates. These four features, in a very similar arrangement, are present in both chymotrypsin and subtilisin even though they are achieved in the two enzymes in completely different ways by quite different three-dimensional structures. Chymotrypsin is built up from two p-barrel domains, whereas the subtilisin structure is of the a/p type. These two enzymes provide an example of convergent evolution where completely different loop regions, attached to different framework structures, form similar active sites. 

The elucidation of the X-ray structure of chymotrypsin (Ref. 1) and in a later stage of subtilisin (Ref. 2) revealed an active site with three crucial groups (Fig. 7.1)-the active serine, a neighboring histidine, and a buried aspartic acid. These three residues are frequently called the catalytic triad, and are designated here as Aspc Hisc Serc (where c indicates a catalytic residue). The identification of the location of the active-site groups and intense biochemical studies led to several mechanistic proposals for the action of serine proteases (see, for example, Refs. 1 and 2). However, it appears that without some way of translating the structural information to reaction-potential surfaces it is hard to discriminate between different alternative mechanisms. Thus it is instructive to use the procedure introduced in previous chapters and to examine the feasibility of different... 

Figure 9-6. Selective proteolysis and associated conformational changes form the active site of chymotrypsin, which includes the Aspl 02-His57-Ser195 catalytic triad. Successive proteolysis forms prochymotrypsin (pro-CT), Jt-chymotrypsin (jt-CT),and ultimately a-chymotrypsin (a-CT), an active protease whose three peptides remain associated by covalent inter-chain disulfide bonds.

NS3 is a 631 amino acid protein, and its first 180 amino acids encode a serine protease of the chymotrypsin family (Figure 2.2A). It has a typical chymotrypsin-family fold consisting of two jS-barrels, with catalytic triad residues at the interface. His-57 and Asp-81 are contributed by the N-terminal jS-barrel and Ser-139 from the C-terminal jS-barrel. NS3 and closely related viral proteases are significantly smaller than other members of the chymotrypsin family, and many of the loops normally found between adjacent jS-strands in trypsin proteases are truncated in NS3 [31]. Probably... 

Fig. 3a-c. Schematic representation of the three residues involved in the formation of the catalytic triad of chymotrypsin [82] with the pseudo-atom vectors (arrows) overlaid a the positions of real non-hydrogen atoms in the sidechains, residues b the S and E pseudo-atom positions with the associated distances between them (dotted lines) (for clarity the MM distances are not shown) c the search matrix for this grouping of sidechains, the distances, all of which are in A, being for the SS, SE, ES and EE distances respectively... 

The catalytic residues of serine proteases such as chymotrypsin generally involve catalytic triad of Asp, His, and Ser residues, for example,... 

Search the Enzyme Structure Database for y-chymotrypsin active site (by the aid of the active-site-modified enzyme or active-site-specific inhibitor-enzyme complex) to identify and depict (save pdb file) the catalytic triad of y-chymotrypsin. 

Perform the regression analyses for the descriptors to assess the contribution of substituent effect(s) on the rate of a-chymotrypsin-catalyzed hydrolysis of p-nit-rophenyl esters. Referring to the catalytic triad of chymotrypsin, rationalize your results for the plausible reaction mechanism. 

A similar concept was used in the development of artificial chymotrypsin mimics [54]. The esterase-site was modeled by using the phosphonate template 75 as a stable transition state analogue (Scheme 13.19). The catalytic triad of the active site of chymotrypsin - that is, serine, histidine and aspartic acid (carboxy-late anion) - was mimicked by imidazole, phenolic hydroxy and carboxyl groups, respectively. The catalytically active MIP catalyst 76 was prepared using free radical polymerization, in the presence of the phosphonate template 75, methacrylic acid, ethylene glycol dimethacrylate and AIBN. The template removal conditions had a decisive influence on the efficiency of the polymer-mediated catalysis, and best results were obtained with aqueous Na2CC>3. 

Amongst the subnanomolar chymotrypsin inhibitors, modelling of one of the best variants implied a novel inhibitory mechanism for protein serine protease inhibitors, in which two amino acid side chains (arginine and aspartic acid) intrude into the proximity of the catalytic triad of the protease rather than binding in the substrate-binding pockets (see Fig. 4). 

---