Trypsin catalyzed hydrolysis and

The peptide shown m green was isolated by trypsin catalyzed hydrolysis and has an ammo acid sequence that completes the remaining overlaps... 

Figure 4. Trypsin-catalyzed hydrolysis and condensation of trimethylethoxysilane at 10° C.

Wang, S.S., and Carpenter, F.H. (1968) Kinetic studies at high pH of the trypsin-catalyzed hydrolysis of Na-benzoyl derivatives of L-arginamide, L-lysinamide, and S-2-aminoethyl-L-cysteinamide and related compounds./. Biol. Chem. 243, 3702-3710. 

The same enzyme was used for the hydrolysis ofp-nitrophenyl-P-D-galactopyrano-side to D-galactose (Scheme 4.97) by Kanno et al. [409,410] in a PMMA microreactor. Quantitative hydrolysis was reported and the reaction was about five times faster than the batch reaction. This unexpected rate enhancement is one of the few examples in which a difference was observed in kinetics between batch and microscale. A second example was reported by Maeda and coworkers [411], They described the trypsin-catalyzed hydrolysis of benzoylarginine-p-nitroanilide and found that the rate of reaction seemed to be 20 times greater than the batchwise system (Scheme 4.98). 

Table 2. Kinetic parameters for trypsin-catalyzed hydrolysis of 8 and 9...

Fig. 3. Time course of the trypsin-catalyzed hydrolysis of p-amidinophenyl acetate (46 R = CH3) at pH 8.0, 25 °C. Concentrations of enzyme and ester are 10 pM and 0.7 mM, respectively...

The reaction process of trypsin-catalyzed hydrolysis of the inverse substrates is illustrated in Fig. 4. Here the process is compared to that of normal-type substrates. After specific binding and efficient acylation, the site-specific amidinophenyl moiety is cleaved (leaving group) to give the acyl enzyme in a very specific manner. As a result, inverse substrates are expected to be applicable as a general method for specific introduction of any acyl group of non-specific structure into the trypsin active site. 

Trypsin-catalyzed hydrolysis gives three peptides and a single amino acid ... 

The trypsin catalyzed hydrolysis (Fig. 7A) of the highly specific substrate Na carbobenzyloxy-L-lysine-ji-nitrophenyl-ester (Z-lys-pNP) has been studied in detail under cryoenzymological conditions by both spectrophotometry and NMR spectroscopy. The kinetic data from both techniques confirm that the kinetics and mechanism under cryoenzymological conditions are essentially the same as those determined at ambient temperatures by rapid reaction techniques. The hydrolysis of [ C]Z-lys-pNP[(S) in Fig. 7A, 8 = 173.6 ppm] by trypsin was monitored by the decrease in intensity of this signal and the increase in the signal arising from the product (P2 in Fig. 7A, 8 = 177.7... 

Figure 5. Turnover numbers of the trypsin-catalyzed hydrolysis of trimethylethoxysilane and condensation of trimethylsilanol at 10 C.

Asaad, N. and Engberts, J.B.F.N., Cytosol-mimetic chemistry kinetics of the trypsin-catalyzed hydrolysis of p nitrophenyl acetate upon addition of polyethylene glycol and n-tert-butyl acetoacetamide, /. Am. Chem. 

Trypsin-catalyzed hydrolysis forms the following two amino acids and two peptides ... 

Partial hydrolysis of a peptide can be carried out either chemically with aqueous acid or enzymatically. Acidic hydrolysis is unselective and leads to a more or less random mixture of small fragments, but enzymatic hydrolysis is quite specific. The enzyme trypsin, for instance, catalyzes hydrolysis of peptides only at the carboxyl side of the basic amino acids arginine and lysine chymotrypsin cleaves only at the carboxyl side of the aryl-substituted amino acids phenylalanine, tyrosine, and tryptophan. 

Fig. 3.3. Major steps in the hydrolase-catalyzed hydrolysis of peptide bonds, taking chymo-trypsin, a serine hydrolase, as the example. Asp102, His57, and Ser195 represent the catalytic triad the NH groups of Ser195 and Gly193 form the oxyanion hole . Steps a-c acylation Steps d-f deacylation. A possible mechanism for peptide bond synthesis by peptidases is represented by the reverse sequence Steps f-a.

More than a third of all known proteolytic enzymes are serine proteases (2). The family name stems from the nucleophilic serine residue within the active site, which attacks the carbonyl moiety of the substrate peptide bond to form an acyl-enzyme intermediate. Nucleophilicity of the catalytic serine is commonly dependent on a catalytic triad of aspartic acid, histidine, and serine—commonly referred to as a charge relay system (3). First observed by Blow over 30 years ago in the structure of chymotrypsin (4), the same combination has been found in four other three-dimensional protein folds that catalyze hydrolysis of peptide bonds. Examples of these folds are observed in trypsin. 

There are no significant differences between ethyl and methyl esters concerning synthesis or cleavage. Related protocols of the methyl esters (see Section 2.2.1.1.1.1) are, therefore, applied to the ethyl esters. The usefulness of ethyl esters is somewhat limited by the difficulties encountered in their saponification. Hydrolysis with alkali is feasible, but ethyl esters are less sensitive to nucleophilic attack than methyl esters. Aminolysis and hydrazinolysis as well as cleavage of the alkyl-oxygen bond with lithium iodide in pyridineb l proceed several times slower in the case of ethyl esters. Mild enzyme-catalyzed hydrolysis by trypsin and chymotrypsin,t 2° 2 1 or by carboxypeptidase remains an attractive alternative. 

Enzyme-catalyzed hydrolysis, exploiting the esterase activity of proteases such as trypsin and chymotrypsint ° l or carboxypeptidase has opened alternative routes to the deprotection of several peptide methyl, ethyl, and ferf-butyl esters. In fact, methyl, ethyl, and benzyl esters are successfully hydrolyzed from protected peptides using the alkaline protease from Bacillus subtilis or alcalase from Bacillus licheniformis which accepts... 

Many enz5mies have been named by adding the suffix "-ase" to the name of their substrate or to a word or phrase describing their activity, e.g. UREASE catalyze hydrolysis of urea, MALTASE act on maltose, and DNA polymerase catalyze the synthesis of DNA. Other enzymes such as PEPSIN and TRYPSIN have names they do not denote their substrates. 

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