Chymotrypsin homologs

Figure 2.19 Organization of polypeptide chains into domains. Small protein molecules like the epidermal growth factor, EGF, comprise only one domain. Others, like the serine proteinase chymotrypsin, are arranged in two domains that are required to form a functional unit (see Chapter 11). Many of the proteins that are involved in blood coagulation and fibrinolysis, such as urokinase, factor IX, and plasminogen, have long polypeptide chains that comprise different combinations of domains homologous to EGF and serine proteinases and, in addition, calcium-binding domains and Kringle domains.

Serine proteinase domains that are homologous to chymotrypsin, which has about 245 amino acids arranged in two domains. 

Leu—CH2CI than Tos—Phe—CH2CI in contrast to chy-motrypsin (note chymotrypsin A and chymotypsin B were later found to be homologous and now are listed with EC 3.4.21.1). 

Sequences have been determined for plasminogen and bovine Factor XII, and they are not homologous with the other serine proteases. The amino-terminal sequence of Factor XII is homologous, however, with the active site of several naturally occurring protease inhibitors (11). Numbering corresponds to chymotrypsin with active serine at 195. 

The mammalian serine proteases appear to represent a classic case of divergent evolution. All were presumably derived from a common ancestral serine protease.23 Proteins derived from a common ancestor are said to be homologous. Some nonmammalian serine proteases are 20 to 50% identical in sequence with their mammalian counterparts. The crystal structure of the elastase-like protease from Streptomyces griseus has two-thirds of the residues in a conformation similar to those in the mammalian enzymes, despite having only 186 amino acids in its sequence, compared with 245 in a-chymotrypsin. The bacterial enzymes and the pancreatic ones have probably evolved from a common precursor. 

For instance, the mammalian serine proteases — trypsin, chymotrypsin, and elastase—are very similar in structure and conformation. If a new mammalian serine protease is discovered, and sequence homology with known proteases... 

Fig. 2. Mapping of conserved features onto three-dimensional structure. The sequence differences between chymotrypsin and its closest homologs are mapped onto a surface representation of the structure of chymotrypsin (PDB lab9). Apeptide ligand (TPGVY) is show in stick format.

Often-cited examples for tandem duplications are pepsin, a member of the aspartate protease family [19], and chymotrypsin, representative of the trypsin-like protease family [20], Structurally, both enzymes consist of two homologous, stacked domains, fi-... 

Sequence 1, in Table I, is the sequence of trypsin, chymotrypsin, pancreatic elastase, thrombin, and other mammalian proteases, and it occurs throughout the animal kingdom down to invertebrates as primitive as the sea anemone (4). The Streptomyces griseus enzymes are from Pronase, a commercial enzyme preparation. Two of its components, Streptomyces griseus trypsin and protease A, not only have the Asp.Ser.Gly sequence, but they show several other homologies in sequence with the mammalian enzymes (5, 6), The same is true for the sequence of a-lytic protease, the Myxobacter 495 enzyme (7). 

The amino acid sequence of the enzyme is homologous with those of the pancreatic enzymes and has been shown by model building to be compatible with a chymotrypsin-like three-dimensional structure and catalytic site (36). The homology in sequence is particularly well marked around His-57. The enzyme s catalytic properties are virtually indistinguishable from those of pancreatic elastase. 

The evolutionary pathways leading to such a scheme are, in part, reflected in the apparent structural homology which exists between these proteases. Sequence information points to a close relationship between trypsin and chymotrypsin (8), and there are marked similarities in the structures of carboxypeptidases A and B, both to each other and to pepsin. Such relationships seem to exist but are beyond the scope of this article. 

Many other proteins have subsequently been found to contain catalytic triads similar to that discovered in chymotrypsin. Some, such as trypsin and elastase, are obvious homologs of chymotrypsin. The sequences of these proteins are approximately 40% identical with that of chymotrypsin, and their overall structures are nearly the same (Figure 9.12). These proteins operate by mechanisms identical with that of chymotrypsin. However, they have very different substrate specificities. Trypsin cleaves at the peptide bond after residues with long, positively charged side chains—namely, arginine and lysine—whereas elastase cleaves at the peptide bond after amino acids with small side chains—such as alanine and serine. Comparison of the Sj pockets of these enzymes reveals the basis of the specificity. 

Yet another example of the catalytic triad has been found in carboxypeptidase II from wheat. The structure of this enzyme is not significantly similar to either chymotrypsin or subtilisin (Figure 9.15). This protein is a member of an intriguing family of homologous proteins that includes esterases such as acetylcholine esterase and certain lipases. These enzymes all make use of histidine-activated nucleophiles, but the nucleophiles may be cysteine rather than serine. 

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