Chymotrypsin (EC 3.4.21.1, chymotrypsins A and B, alpha-chymar ophth, avazyme, chymar, chymotest, enzeon, quimar, quimotrase, alpha-chymar, alpha-chymotrypsin A, alpha-chymotrypsin) is a digestive enzyme component of pancreatic juice acting in the duodenum, where it performs proteolysis, the breakdown of proteins and polypeptides.[2] Chymotrypsin preferentially cleaves peptide amide bonds where the side chain of the amino acid N-terminal to the scissile amide bond (the P1 position) is a large hydrophobic amino acid (tyrosine, tryptophan, and phenylalanine).[3] These amino acids contain an aromatic ring in their side chain that fits into a hydrophobic pocket (the S1 position) of the enzyme. It is activated in the presence of trypsin. The hydrophobic and shape complementarity between the peptide substrate P1 side chain and the enzyme S1 binding cavity accounts for the substrate specificity of this enzyme.[4][5] Chymotrypsin also hydrolyzes other amide bonds in peptides at slower rates, particularly those containing leucine at the P1 position.[3]
| Chymotrypsin | |||||||||
|---|---|---|---|---|---|---|---|---|---|
 Crystallographic structure of Bos taurus chymotrypsinogen[1] | |||||||||
| Identifiers | |||||||||
| EC no. | 3.4.21.1 | ||||||||
| CAS no. | 9004-07-3 | ||||||||
| Databases | |||||||||
| IntEnz | IntEnz view | ||||||||
| BRENDA | BRENDA entry | ||||||||
| ExPASy | NiceZyme view | ||||||||
| KEGG | KEGG entry | ||||||||
| MetaCyc | metabolic pathway | ||||||||
| PRIAM | profile | ||||||||
| PDB structures | RCSB PDB PDBe PDBsum | ||||||||
| Gene Ontology | AmiGO / QuickGO | ||||||||
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| Chymotrypsin C | |||||||||
|---|---|---|---|---|---|---|---|---|---|
| Identifiers | |||||||||
| EC no. | 3.4.21.2 | ||||||||
| CAS no. | 9036-09-3 | ||||||||
| Databases | |||||||||
| IntEnz | IntEnz view | ||||||||
| BRENDA | BRENDA entry | ||||||||
| ExPASy | NiceZyme view | ||||||||
| KEGG | KEGG entry | ||||||||
| MetaCyc | metabolic pathway | ||||||||
| PRIAM | profile | ||||||||
| PDB structures | RCSB PDB PDBe PDBsum | ||||||||
| |||||||||
Structurally, it is the archetypal structure for its superfamily, the PA clan of proteases.
Activation
editChymotrypsin is synthesized in the pancreas. Its precursor is chymotrypsinogen. Trypsin activates chymotrypsinogen by cleaving peptidic bonds in positions Arg15 – Ile16 and produces π-chymotrypsin. In turn, aminic group (-NH3+) of the Ile16 residue interacts with the side chain of Asp194, producing the "oxyanion hole" and the hydrophobic "S1 pocket". Moreover, chymotrypsin induces its own activation by cleaving in positions 14–15, 146–147, and 148–149, producing α-chymotrypsin (which is more active and stable than π-chymotrypsin).[6] The resulting molecule is a three-polypeptide molecule interconnected via disulfide bonds.
Mechanism of action and kinetics
editThis article is missing information about which specific protein this residue numbering is based on (use hatnote). (June 2022) |
In vivo, chymotrypsin is a proteolytic enzyme (serine protease) acting in the digestive systems of many organisms. It facilitates the cleavage of peptide bonds by a hydrolysis reaction, which despite being thermodynamically favorable, occurs extremely slowly in the absence of a catalyst. The main substrates of chymotrypsin are peptide bonds in which the amino acid N-terminal to the bond is a tryptophan, tyrosine, phenylalanine, or leucine. Like many proteases, chymotrypsin also hydrolyses amide bonds in vitro, a virtue that enabled the use of substrate analogs such as N-acetyl-L-phenylalanine p-nitrophenyl amide for enzyme assays.
Chymotrypsin cleaves peptide bonds by attacking the unreactive carbonyl group with a powerful nucleophile, the serine 195 residue located in the active site of the enzyme, which briefly becomes covalently bonded to the substrate, forming an enzyme-substrate intermediate. Along with histidine 57 and aspartic acid 102, this serine residue constitutes the catalytic triad of the active site. These findings rely on inhibition assays and the study of the kinetics of cleavage of the aforementioned substrate, exploiting the fact that the enzyme-substrate intermediate p-nitrophenolate has a yellow colour, enabling measurement of its concentration by measuring light absorbance at 410 nm.
Chymotrypsin catalysis of the hydrolysis of a protein substrate (in red) is performed in two steps. First, the nucleophilicity of Ser-195 is enhanced by general-base catalysis in which the proton of the serine hydroxyl group is transferred to the imidazole moiety of His-57 during its attack on the electron-deficient carbonyl carbon of the protein-substrate main chain (k1 step). This occurs via the concerted action of the three-amino-acid residues in the catalytic triad. The buildup of negative charge on the resultant tetrahedral intermediate is stabilized in the enzyme's active site's oxyanion hole, by formation of two hydrogen bonds to adjacent main-chain amide-hydrogens.
The His-57 imidazolium moiety formed in the k1 step is a general acid catalyst for the k-1 reaction. However, evidence for similar general-acid catalysis of the k2 reaction (Tet2)[7] has been controverted;[8] apparently water provides a proton to the amine leaving group.
Breakdown of Tet1 (via k3) generates an acyl enzyme, which is hydrolyzed with His-57 acting as a general base (kH2O) in formation of a tetrahedral intermediate, that breaks down to regenerate the serine hydroxyl moiety, as well as the protein fragment with the newly formed carboxyl terminus.
Uses
editMedical uses
editChymotrypsin has been used during cataract surgery.[9] It was marketed under the brand name Zolyse.[10]
Isozymes
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See also
editReferences
edit- ^ PDB: 1CHG; Freer ST, Kraut J, Robertus JD, Wright HT, Xuong NH (April 1970). "Chymotrypsinogen: 2.5-angstrom crystal structure, comparison with alpha-chymotrypsin, and implications for zymogen activation". Biochemistry. 9 (9): 1997–2009. doi:10.1021/bi00811a022. PMID 5442169.
- ^ Wilcox PE (1970). "[5] Chymotrypsinogens—chymotrypsins". Chymotrypsinogens — chymotrypsins. Methods in Enzymology. Vol. 19. pp. 64–108. doi:10.1016/0076-6879(70)19007-0. ISBN 978-0-12-181881-4.
- ^ a b Cotten, Steven W. (2020-01-01), Clarke, William; Marzinke, Mark A. (eds.), "Chapter 33 - Evaluation of exocrine pancreatic function", Contemporary Practice in Clinical Chemistry (Fourth Edition), Academic Press, pp. 573–585, ISBN 978-0-12-815499-1, retrieved 2023-03-18
- ^ Appel W (December 1986). "Chymotrypsin: molecular and catalytic properties". Clin. Biochem. 19 (6): 317–22. doi:10.1016/S0009-9120(86)80002-9. PMID 3555886.
- ^ Berger A, Schechter I (February 1970). "Mapping the active site of papain with the aid of peptide substrates and inhibitors". Philos. Trans. R. Soc. Lond. B Biol. Sci. 257 (813): 249–64. Bibcode:1970RSPTB.257..249B. doi:10.1098/rstb.1970.0024. PMID 4399049. S2CID 6877875.
- ^ Phillips, Jo (2019). Fundamentals of Enzymology. EDTECH. p. 117. ISBN 9781839471605.
- ^ Fersht, A.R.; Requena, Y. (1971). "Mechanism of the -Chymotrypsin-Catalyzed Hydrolysis of Amides. pH Dependence of kc and Km". J. Am. Chem. Soc. 93 (25): 7079–87. doi:10.1021/ja00754a066. PMID 5133099.
- ^ Zeeberg, B.; Caswell, M.; Caplow, M. (1973). "Concerning a reported change in rate-determining step in chymotrypsin catalysis". J. Am. Chem. Soc. 95 (8): 2734–5. doi:10.1021/ja00789a081. PMID 4694533.
- ^ REED H (April 1960). "Chymotrypsin in cataract surgery". Canadian Medical Association Journal. 82 (15): 767–70. PMC 1938008. PMID 14436866.
- ^ "Final List of Withdrawn Applications for Biological Products That Were Removed From the Orange Book on 3-23-20 | FDA". www.fda.gov. Retrieved 5 April 2024.
Further reading
edit- Stryer L, Berg JM, Tymoczko JL (2002). Biochemistry. San Francisco: W.H. Freeman. ISBN 0-7167-4684-0.
- Grisham CM, Reginald H (2005). Biochemistry. Australia: Thomson Brooks/Cole. ISBN 0-534-49033-6.
External links
edit- The MEROPS online database for peptidases and their inhibitors: S01.001 Archived 2019-09-15 at the Wayback Machine
- Chymotrypsin at the U.S. National Library of Medicine Medical Subject Headings (MeSH)
