Protein hydrolysates aminopeptidase

Aminopeptidase Other hydrolases Mold, bacteria Debittering of protein hydrolysates... 

Methods for eliminating bitter peptides in partial protein hydrolysates are known, but they cause a significant loss of essential amino acids. These procedures usually include additional enzymatic hydrolysis under controlled conditions (a shorter time for the hydrolysis leads to higher peptides that are not bitter) and a selection of suitable proteases, such as aminopeptidases, carboxypeptidases and some other proteases. Enzymes of plant and microbial origin have been successfully used for this purpose. For example, the intracellular peptidases from Lactococcus lactis ssp. cremoris and Brevibacterium linens, which have high proteolytic activity, successfully hydrolyse bitter peptides in cheeses. 

Procedure 3 papain, leucine aminopeptidase, prolidase This early procedure for enzymic hydrolysis of proteins was reported by Hill and Schmidt (1962) to be successful for hydrolysis of several proteins. Papain was found to be superior to subtilisin or a combination of trypsin and chymotrypsin for the initial hydrolysis. The method might be improved if aminopeptidase M (discovered after the method was developed) is used in place of the leucine aminopeptidase, but to our knowledge this has not been tested. The problem with diketo-piperazine formation from X-Pro dipeptides in aminopeptidase M hydrolysates of peptides (see above) may make this substitution less desirable than it would seem at first. 

A limited amount of information can be obtained by the use of proteolytic enzymes that detach either amino acids or dipeptides sequentially from the C-terminus. They are thus complementary to the aminopeptidases and dipeptidyl aminopeptidases. Two pancreatic enzymes, carboxypeptidases A and B, differ in specificity. The former preferentially liberates C-terminal amino acids with aromatic side chains, somewhat less readily amino acids with alkyl side chains and, more slowly still, other amino acids, but not Pro, Arg, Lys and His. In contrast, carboxypeptidase B releases only C-terminal Arg, Lys and His. Carboxypeptidase Y is much less specific and is capable of removing all amino acids, although Gly and Pro are liberated only slowly. As with aminopeptidases, it is advisable to analyse the hydrolysate at intervals in order to determine the C-terminal sequence of amino acids. An interesting recent development (Carles et al., 1988) uses carboxypeptidase to effect transpeptidation between the protein being sequenced and a tritiated amino acid. The labelled protein is then degraded by various specific methods and then the labelled fragments are isolated by gel electrophoresis and subjected to Edman degradation. 

The oligopeptides formed by the action of the endopeptidases are broken down into their constituent amino acids by the action of the exopeptidases. The carboxypeptidase of the pancreas splits amino acids one by one from the C-terminus so that, by the time they reach the absorbing cells of the small intestine, the dietary proteins have been converted into a mixture of amino acids and small peptides. The mucosal cells which contain both aminopeptidases and dipeptidase take up the small peptides which are then hydrolysed either within the brush border or in the layer immediately beneath it. Thus the final stages of protein digestion, like those of carbohydrates, are intracellular. Under normal circumstances no peptides pass across the mucosa to enter the bloodstream. 

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