α-Chymotrypsin Property

Melting point:

127°C

Density 

1.37[at 20℃]

vapor pressure 

0Pa at 25℃

storage temp. 

-20°C

solubility 

Reconstitute in 1mM HCl. Soluble at 10mg/ml in 1mM HCl. 2mM calcium chloride serves as a stabilizer. Store aliquoted solutions at -20°C for up to a week.

form 

salt-free, lyophilized powder

color 

white

Water Solubility 

125g/L at 25℃

Merck 

13,2282

LogP

-1.3 at 20℃

EPA Substance Registry System

Chymotrypsin (9004-07-3)

Safety

Hazard Codes 

Xn,B

Risk Statements 

36/37/38-42/43-42

Safety Statements 

26-36-36/37-24-22

WGK Germany 

3

RTECS 

GC3050000

F 

3-10

TSCA 

Yes

HS Code 

35079090

Hazardous Substances Data

9004-07-3(Hazardous Substances Data)

Hazard and Precautionary Statements (GHS)

Symbol(GHS)

Signal word

Danger

Hazard statements

H315Causes skin irritation

H317May cause an allergic skin reaction

H319Causes serious eye irritation

H334May cause allergy or asthma symptoms or breathing difficulties if inhaled

H335May cause respiratory irritation

Precautionary statements

P280Wear protective gloves/protective clothing/eye protection/face protection.

P302+P352IF ON SKIN: wash with plenty of soap and water.

P305+P351+P338IF IN EYES: Rinse cautiously with water for several minutes. Remove contact lenses, if present and easy to do. Continuerinsing.

α-Chymotrypsin Chemical Properties,Usage,Production

Degradation

The rate and extent of insulin degradation by trypsin and α-Chymotrypsin were examined in vitro, and the initial sites of cleavage by α-Chymotrypsin were identified. The apparent Km for both enzymes was approximately the same, but the apparent Vmax for α-Chymotrypsin was 8.6 times greater. At a molar ratio of 172:1 (insulin: enzyme), chymotrypsin caused near-total loss of insulin within 40 min, while very little insulin was degraded by trypsin. Chymotrypsin appeared to cleave initially at the carboxyl side of the B26-Tyr and A19-Tyr residues, and additional cleavage at the B16-Tyr, B25-Phe, and A14-Tyr residue sites also occurred rapidly. Only two to three other susceptible bonds, which are not exposed at the surface of the insulin molecule, remained intact after the quenching of initial cleavage[1].

Chemical Properties

Lyophilized powder, dialyzed

Uses

α-Chymotrypsin from bovine has been used in a study to inform proteasome inhibition in order to advance anticancer research. α-Chymotrypsin from bovine has also been used in a study that functionalized surface anchored poly(methylhydrosiloxane) thin films on oxidized silicon wafers.

Uses

The enzyme from Sigma has been used to assess the effect of limited proteolysis with α-chymotrypsin on the sperm penetration.

Uses

α-Chymotrypsin from bovine pancreas has been used in a study to investigate protein extraction by Winsor-III microemulsion systems. α-Chymotrypsin from bovine pancreas has also been used in a study to investigate a new specific fullerene-based fluorescent probe for trypsin.

General Description

Chymotrypsin (Chymar) is extractedfrom mammalian pancreas and is used in cataractsurgery. A dilute solution is used to irrigate the posteriorchamber of the eye to dissolve the fine filaments that holdthe lens.

Biochem/physiol Actions

α-Chymotrypsin is a serine peptidase and has 241 amino acid residues contained in three polypeptide chains (A chain-13 residues, B chain-131 residues, and C chain-97 residues) linked by disulfide bridges. Molecular weight of this enzyme is found to be 25 kDa. Its pI is 8.75. It selectively hydrolyzes peptide bonds on the C-terminal side of tyrosine, phenylalanine, tryptophan, and leucine. Ca2+ activates and stabilizes the enzyme. The enzyme is inhibited by diisopropyl fluorophosphate (DFP), phenylmethanesulfonyl fluoride (PMSF), N-p-tosyl-L-phenylalanine chloromethyl ketone (TPCK), chymostatin, aprotinin, α1-antitrypsin, α2-macroglobulin, 10 mM Cu2+ and Hg2+.

Mechanism of action

The general mechanism for chymotrypsin is the classic serine protease mechanism. Hydrolytic proteolysis by α-chymotrypsin begins with an initial nucleophilic attack on the peptide bond by Ser 195, activated by deprotonation by His 57. This leads to forming a tetrahedral intermediate, stabilized by the amide groups of Ser 195 and Gly 193. The subsequent collapse of this intermediate, assisted by protonation of the leaving group by His 57 and Asp 102, leads to an acyl-enzyme intermediate. Activation of a water molecule by His 57 and Asp 102 facilitates hydrolysis of this intermediate, resulting in the reformation of the catalytically active serine residue and the release of the product facilitated by protonation with His 57.

Purification Methods

α-Chymotrypsin is crystallised twice from four-tenths saturated ammonium sulfate solution, then dissolved in 1mM HCl and dialysed against 1mM HCl at 2-4o. The solution is stored at 2o [Lang et al. J Am Chem Soc 80 4923 1958].

References

[1] R J Schilling, A K Mitra. “Degradation of insulin by trypsin and alpha-chymotrypsin.” Pharmaceutical Research 8 6 (1991): 721–7.