Chymotrypsin

chymotrypsin
Crystallographic structure of Bos taurus chymotrypsinogen.[1]
Identifiers
EC number	3.4.21.1
CAS number	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 / EGO
Search PMC articles PubMed articles

chymotrypsin C
Identifiers
EC number	3.4.21.2
CAS number	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
Search PMC articles PubMed articles

Chymotrypsin is a digestive enzyme that can perform proteolysis.^[2] Chymotrypsin preferentially cleaves peptide amide bonds where the carboxyl side of the amide bond (the P₁ position) is a tyrosine, tryptophan, or phenylalanine. These amino acids contain an aromatic ring in their sidechain that fits into a 'hydrophobic pocket' (the S₁ position) of the enzyme. The hydrophobic and shape complementarity between the peptide substrate P₁ sidechain and the enzyme S₁ binding cavity accounts for the substrate specificity of this enzyme.^[3][4] Chymotrypsin also hydrolyzes other amide bonds in peptides at slower rates, particularly those containing leucine at the P₁ position.

Activating chymotrypsinogen into chymotrypsin

Chymotrypsin is synthesized in the pancreas by protein biosynthesis as a precursor called chymotrypsinogen that is enzymatically inactive. On cleavage by trypsin into two parts that are still connected via an S-S bond, cleaved chymotrypsinogen molecules can activate each other by removing two small peptides in a trans-proteolysis. The resulting molecule is active chymotrypsin, a three-polypeptide molecule interconnected via disulfide bonds.

Action and kinetics of chymotrypsin

In vivo, chymotrypsin is a proteolytic enzyme (Serine protease) acting in the digestive systems of mammals and other organisms. It facilitates the cleavage of peptide bonds by a hydrolysis reaction, which despite being thermodynamically favourable occurs extremely slowly in the absence of a catalyst. The main substrates of chymotrypsin include tryptophan, tyrosine, phenylalanine, leucine, and methionine, which are cleaved at the carboxyl terminal. Like many proteases, chymotrypsin will also hydrolyse 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.

Mechanism of peptide bond cleavage in α-chymotrypsin

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.

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 us to measure its concentration by measuring light absorbance at 410 nm.

It was found that the reaction of chymotrypsin with its substrate takes place in two stages, an initial “burst” phase at the beginning of the reaction and a steady-state phase following Michaelis-Menten kinetics. It is also called "ping-pong" mechanism. The mode of action of chymotrypsin explains this as hydrolysis takes place in two steps. First acylation of the substrate to form an acyl-enzyme intermediate and then deacylation in order to return the enzyme to its original state.

Isozymes

chymotrypsinogen B1 Identifiers Symbol CTRB1 Entrez 1504 HUGO 2521 OMIM 118890 RefSeq NM_001906 UniProt P17538 Other data EC number 3.3.21.1 Locus Chr. 16 q23.1

chymotrypsinogen B2 Identifiers Symbol CTRB2 Entrez 440387 HUGO 2522 RefSeq NM_001025200 UniProt Q6GPI1 Other data EC number 3.3.21.1 Locus Chr. 16 q22.3

chymotrypsin C (caldecrin) Identifiers Symbol CTRC Entrez 11330 HUGO 2523 OMIM 601405 RefSeq NM_007272 UniProt Q99895 Other data EC number 3.3.21.2 Locus Chr. 1 p36.21

References

1. ^ 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. 
2. ^ Wilcox PE (1970). "Chymotrypsinogens — chymotrypsins". Methods in Enzymology. Methods in Enzymology 19: 64–108. doi:10.1016/0076-6879(70)19007-0. ISBN 978-0-12-181881-4. 
3. ^ 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. 
4. ^ 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. doi:10.1098/rstb.1970.0024. PMID 4399049. 

Further reading

• 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

• The MEROPS online database for peptidases and their inhibitors: S01.001
• MeSH Chymotrypsin