Trypsin protein-derivatives hydrolysis

A number of alkylating agents yield derivatives which are stable under the conditions for acidic hydrolysis of proteins. The reaction with ethylene imine giving an S-aminoethyl derivative and, hence, an additional linkage position in the protein for hydrolysis by trypsin, was mentioned in Section 1.4.1.3. lodoacetic acid, depending on the pH, can react with cysteine, methionine, lysine and histidine residues ... 

Hydrolysis of peptides and proteins in the GI tract can occur luminally, at the brash border and intracellularly. Luminal activity from the pancreatic proteases trypsin, chymotrypsin, elastase and carboxypeptidase A is mainly directed against large dietary proteins. The main enzymatic activity against small bioactive peptides is derived from the bmsh border of the enterocyte. Brash border proteases, such as aminopeptidase A and N, diaminopeptidease IV and Zn-stable Asp-Lys peptidase, preferentially cleave oligopeptides of up to 10 ammo acid residues and are particularly effective in the cleavage of tri- and tetra-peptides. 

The specificity of chymotrypsin for hydrolysis of peptide bonds formed by the carbo,xyl groups of tyrosine, phenylalanine, and tryptophan has been recognized for some time (Green and Neurath, 1954 Desnuelle, 1960). Action on synthetic substrates of leucine (Goldenberg et al., 1951) and methionine (Kaufman and Neurath, 1949) also has been noted although at much slower rates than observed with the aromatic amino acid derivatives. When protein substrates or synthetic ester substrates are examined, it is evident that a variety of bonds can be hydrolyzed by chymotrypsin. Inagami and Sturtevant (1960) observed that chymotryptic hydrolysis of a-benzoyl-L-arginine ethyl ester, a typical trypsin substrate, occurred at a maximum rate which was 20% of that observed with trypsin. Several ester substrates, such as p-nitrophenylacetate (Hartley and Kilby, 1954), are also hydrolyzed. 

Crafting a new breakpoint. Ethyleneimine reacts with cysteine side chains in proteins to form -aminoethyl derivatives. The peptide bonds on the carboxyl side of these modified cysteine residues are susceptible to hydrolysis by trypsin. Why ... 

Modification of specific amino acid residues should affect the rate of proteolysis by those enzymes which have specificity for the unmodified residues. For example, modification of the lysine residues of a protein restricts the action of trypsin. Dimethylated casein undergoes a slower rate of hydrolysis by trypsin in vitro, and the extent of proteolysis is lower than that of casein (267). An unexpected finding was that the rate and extent of hydrolysis of dimethylated casein by a-chymotrypsin are also adversely affected, while hydrolysis by subtilisin is not. Peptides derived from dimethylated casein were inhibitory of a-chymotrypsin. 

The partly digested material passes next into the small intestine where the process of digestion is carried on by the substances present in the intestinal juice and the pancreatic juice. The latter contains amylopsin and trypsinogen. When the latter comes into contact with the enzyme enterokinase of the intestinal juice trypsin is formed. This enzyme, in the presence of the alkali in the intestinal juice, causes the hydrolysis of all forms of proteins. Unlike pepsin it causes the hydrolysis to proceed essentially to the complete splitting of the molecule to amino-acids. The juice secreted by the small intestine contains the proteolytic enzyme erepsin, which, with a few exceptions, induces the hydrolysis of only the derived proteins to amino-acids. 

Fig. 2.13 Dilatational rheology parameters derived Irom oscillation of a pendant droplet during a frequency sweep of p-lactoglobulin and its hydrolysates at the MCT oil-water interface, protein content 0.01 wt%, pH 8.0, amplitude AA/A = 2.8 %. Hydrolysates have been produced using trypsin (T) or alcalase (A). Percentage in sample names indicate the degree of hydrolysis...

A novel mass spectrometric method has been described that permits the identification of the C-terminal peptide of a protein (68). The technique involves the incorporation of into all a-C02H groups released during enzyme-catalysed partial hydrolysis e. g. with trypsin) of the protein in 0-enriched water ( 0 0 50 50). Individual peptides were analyzed by GC/EIMS as their trifluoroacetyl permethyl derivatives and the C-terminal peptide was characterized as the one that did not incorporate (The non-... 

The names of enzymes describe the compound or the reaction that is catalyzed. The actual names of enzymes are derived by replacing the end of the name of the reaction or reacting compound with the suffix ase. For example, an oxidase catalyzes an oxidation reaction, and a dehydrogenase removes hydrogen atoms. The compound sucrose is hydrolyzed by the enzyme sucrase, and a lipid is hydrolyzed by a lipase. Some early known enzymes use names that end in the suffix in, such as papain found in papaya, rennin found in milk, and pepsin and trypsin, enzymes that catalyze the hydrolysis of proteins. 

In order to verify this assumption, we studied the temperature dependence of the kinetic isotope effect during hydrolysis of some specific substrates, i.e. amino acid esters, by serine proteases like a-chymotrypsin[468,469] and 3-trypsin[470]. It is well known that a two-stage hydrolysis mechanism takes place for this class of enzymes with the formation, as an intermediate product, of an acyl derivative of the amino acid serine (Ser-195), a constituent of the polypeptide chain of the protein. The side group (-CH2OH) of serine is located in the region of the active center[471-474] ... 

In the last few years a further type of modified flavocoenzyme has beeen discovered which is structurally related to the coenzymes of succinate dehydrogenase and monoamine oxidase, but differs considerably in its chemical properties. Early literature reports indicated the presence of a flavin in cytochrome C552 from Chromatium which could not be extracted with trichloroacetic acid or acidic ammonium sulfate (7), but could be released, for example by trypsin digestion or by incubation with saturated urea solutions (2). Absorption, fluorescence and ESR behaviour were closely similar to those of 8a-cysteinyl-ribo-flavin (12) and indicated the presence of a covalent link to the protein, through position 8a (185). Strong acid hydrolysis of these peptides liberated the flavin as mixture of riboflavin derivatives oxidation with per-formic acid and acid dephosphorylation yielded a homogeneous riboflavin derivative, which was identical with 8-nor-8-carboxy-riboflavin (185). 

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