Difference between .alpha.- and .delta.-chymotrypsins. Preparation and alkaline pH dependence of .alpha.1-chymotrypsin-catalyzed hydrolysis of N-acetyl-L-tryptophan methyl ester (ATME). Involvement of alanine-149 in .alpha.-chymotrypsin catalysis

1971 ◽  
Vol 93 (15) ◽  
pp. 3783-3784 ◽  
Author(s):  
Myron L. Bender ◽  
Pablo Valenzuela
Keyword(s):  
Methyl Ester ◽  
Ph Dependence ◽  
Alkaline Ph ◽  
Hydrolysis Of ◽  
Tryptophan Methyl Ester

scholarly journals Comparative studies of the specificities of α-chymotrypsin and subtilisin BPN′. Studies with flexible substrates

1972 ◽  
Vol 126 (3) ◽  
pp. 645-657 ◽  
Author(s):  
T. N. Pattabiraman ◽  
W. B. Lawson
Keyword(s):  
Methyl Ester ◽  
Binding Site ◽  
Comparative Studies ◽  
Detrimental Effect ◽  
Flexible Substrates ◽  
Enhancing Effect ◽  
Wide Range ◽  
Phenylalanine Methyl Ester ◽  
Hydrolysis Of ◽  
Tryptophan Methyl Ester

A series of arylalkanoate esters and α-acetamidoarylalkanoate esters were tested as substrates for α-chymotrypsin and subtilisin BPN′. Chymotrypsin hydrolysed N-acetyl-l-phenylalanine methyl ester and methyl 4-phenylbutyrate faster than their respective higher and lower homologues, whereas methyl 2-acetamido-6-phenylhexanoate and methyl 6-phenylhexanoate were better substrates for subtilisin than their lower homologues. N-Acetyl-l-tryptophan methyl ester and its analogue, N-acetyl-3-(1-naphthyl)-alanine methyl ester, were hydrolysed 23 times faster by chymotrypsin than by subtilisin. These results indicate that the binding site of α-chymotrypsin is roughly 1.1nm (11Å) long and curved, whereas that of subtilisin is a longer system and less curved. The stereo-specificity during the hydrolysis of typical substrates by both enzymes was found to vary over a wide range. The enhancing effect of the α-acetamido group in the l-series of substrates and the detrimental effect in the d-series of substrates also varies considerably.

scholarly journals Reinvestigation of the reaction of chymotrypsin with N-furylacryloyltryptophan derivatives at acidic pH

1979 ◽  
Vol 181 (3) ◽  
pp. 733-736 ◽  
Author(s):  
A L Fink ◽  
R Feldman ◽  
J Zehnder
Keyword(s):  
Methyl Ester ◽  
Wide Temperature Range ◽  
Methyl Esters ◽  
Ph Dependence ◽  
Gel Filtration ◽  
Low Ph ◽  
Time Dependent ◽  
Acidic Ph ◽  
Spectral Changes ◽  
Tryptophan Methyl Ester

The reaction of alpha-chymotrypsin with N alpha-3-(2-furyl)acryloyl-L-tryptophan methyl ester (FA-Trp-OMe) and amide has been investigated in aqueous and dimethylsulphoxide cryosolvent solutions from pH2 to 7 and over a wide temperature range. Previous reports have suggested that an intermediate preceding the acyl-enzyme can be detected spectrophotometrically in the reaction with methyl esters of FA-Trp and FA-Tyr at low pH [Yu & Viswanatha (1969) Eur. J. Biochem. 11, 347–352), and that this intermediate is an oxazolinone [Coletti-Previero et al. (1970) FEBS Lett. 11, 213–217]. We show that the previous interpretations of the time-dependent spectral changes were incorrect, and that the only detected intermediate is the acyl-enzyme. This may be isolated by gel filtration at pH less than 2.5, 1 degree C, owing to its relative stability. The pH-dependence of the rates of acylation and deacylation from pH 8.5 to 2.0 are consistent with a single ionization of pK congruent to 7.0 in both aqueous and cryosolvent solutions.

pH dependence of the dephosphorylated pepsin-catalyzed hydrolysis of N-acetyl-L-phenylalanyl-L-tyrosine methyl ester

1970 ◽  
Vol 92 (1) ◽  
pp. 186-189 ◽  
Author(s):  
Gerald E. Clement ◽  
James. Rooney ◽  
David. Zakheim ◽  
James. Eastman
Keyword(s):  
Methyl Ester ◽  
Ph Dependence ◽  
Hydrolysis Of

scholarly journals The structure and mechanism of stem bromelain. Evaluation of the homogeneity of purified stem bromelain, determination of the molecular weight and kinetic analysis of the bromelain-catalysed hydrolysis of N-benzyloxycarbonyl-l-phenylalanyl-l-serine methyl ester

1974 ◽  
Vol 143 (3) ◽  
pp. 575-586 ◽  
Author(s):  
Christopher W. Wharton
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Methyl Ester ◽  
Gel Electrophoresis ◽  
Ph Dependence ◽  
Polyacrylamide Gel Electrophoresis ◽  
Terminal Amino Acid ◽  
Terminal Amino ◽  
Stem Bromelain ◽  
Hydrolysis Of

1. Purified stem bromelain (EC 3.4.22.4) was eluted from Sephadex G-100 as a single peak. The specific activity across the elution peak was approximately constant towards p-nitrophenyl hippurate but increased with elution volume with N2-benzoyl-l-arginine ethyl ester as substrate. 2. The apparent molecular weight, determined by elution analysis on Sephadex G-100, is 22500±1500, an anomalously low value. 3. Purified stem bromelain was eluted from CM-cellulose CM-32 as a single peak and behaved as a single species during column electrophoresis on Sephadex G-100. 4. Purified stem bromelain migrates as a single band during polyacrylamide-gel electrophoresis under a wide variety of conditions. 5. The molecular weight determined by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate is 28500±1000. 6. Sedimentation-velocity and equilibrium-ultracentrifugation experiments, under a variety of conditions, indicate that bromelain is an apparently homogeneous single peptide chain of mol.wt. 28400±1400. 7. The N-terminal amino acid composition is 0.64±0.04mol of valine and 0.36±0.04mol of alanine per mol of enzyme of mol.wt. 28500. (The amino acid recovery of the cyanate N-terminal amino acid analysis was standardized by inclusion of carbamoyl-norleucine at the cyclization stage.) 8. The pH-dependence of the Michaelis parameters of the bromelain-catalysed hydrolysis of N-benzyloxycarbonyl-l-phenylalanyl-l-serine methyl ester was determined. 9. The magnitude and pH-dependence of the Michaelis parameters have been interpreted in terms of the mechanism of the enzyme. 10. The enzyme is able to bind N-benzyloxycarbonyl-l-phenylalanyl-l-serine methyl ester relatively strongly but seems unable to make use of the binding energy to promote catalysis.

Variation in Quantity of Methanol Recovered from Raisins

1963 ◽  
Vol 46 (2) ◽  
pp. 341-343
Author(s):  
M Alice Brown ◽  
James R Woodward ◽  
Floyd DeEds
Keyword(s):  
Methyl Ester ◽  
Esterase Activity ◽  
Enzymic Hydrolysis ◽  
Methanol Content ◽  
Naturally Occurring ◽  
Pectin Esterase ◽  
Hydrolysis Of

Abstract The amount of naturally occurring methanol in fruit must be known so that the quantity left as fumigation residue can be determined. In a study of methanol content of raisins, which had given inconsistent results, the raisins were subjected to different conditions of treatment immediately prior to methanol determination. Conditions that favored pectin esterase activity gave higher values for methanol content than conditions known to inactivate enzymes. Evidence was also obtained that both chemical and enzymic hydrolysis of methyl ester groups of pectic materials occur during analysis.

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