Nonspecific enzymatic hydrolysis

Dipeptidyl peptidase IV (DPP IV, EC 3.4.14.5, CD26) is a serine protease cleaving off dipeptides from the amino terminus of peptides or proteins having proline or alanine at the penultimate position. Since prolylamides are known to play a critical role in peptide structure and function, and because of their high resistance toward nonspecific enzymatic hydrolysis, a few enzymes capable of cleaving this structural motif have attracted consid-... 

Overcoming the problems of acid hydrolysis, scientists have adopted different enzymatic hydrolysis methods using different types of enzymes for the preparation of COS. The polysaccharide structure of the chitin or chitosan is converted to oligosaccharide form by specific enzymatic depolymerization or nonspecific enzymatic depolymerization. 

Ragan and Meyer (354) have recently reported hyaluronic acid contents and relative viscosities of synovial fluids from the knee joints of 35 patients with rheumatoid arthritis and 11 patients without joint diseases. They found with very few exceptions that the apparent degree of polymerization of the hyaluronic acid was lower in the arthritic patients, although the total amount of the mucopolysaccharide present was greater. Hyaluronidase was not detected in joint fluid or in synovial or periarticular tissue and, in any event, its presence would not explain the increase in total hyaluronic acid. Therefore, it was concluded that the defect lies not in an enzymatic hydrolysis or nonspecific depolymerization but in abnormal synthesis of this polysaccharide, involving an increased production of incompletely polymerized hyaluronic acid. 

The azo-BSA release from the microspheres is induced by the hydrolysis of the fatty acid ester side chains on the emulsan. Candida rugosa lipase was used to follow the release of azo-BSA by specific enzymatic cleavage of the fatty acids, and the nonspecific enzymatic cleavage of azo-BSA into sulfanilic acid. The results are displayed in Tabie I. 

In addition to hydrogen ions, other species can also affect the enzymatic catalytic activity. This phenomenon is called inhibition it may be specific, nonspecific, reversible, or irreversible. The inhibition reactions can also be used for the sensing of inhibitors. The best-known example is the sensor for detection of nerve gases. These compounds inhibit the hydrolysis of the acetylcholine ester which is catalyzed by the enzyme acetylcholine esterase. Acetylcholine ester is a key component in the neurotransmission mechanism. 

Two types of T4 lysozyme bindings to cell walls are possible (1) nonspecific attachment and (2) chemical binding associated with the hydrolysis reaction. Events indicated by the fluorescence intensity dropping to background level are associated with T4 lysozyme diffusing away from the cell wall. When the T4 lysozyme attached to the cell wall, many enzymatic reaction turnovers likely occurred in experiments, we have observed that the cell wall typically shrinks and eventually disappears from the imaging field of view [12]. 

That the inactivation was active-site directed was also established in several ways. As mentioned above, the pA a values of k2 and k, were consistent with the pKa value of catalytic activity for a serine protease. Difference spectra of enzyme with inhibitor showed the reactive product being formed in the presence of enzyme. Rates of inhibition decreased in the presence of a known competitive inhibitor, elastatinal (Okura et al., 1975). The reactive intermediate was generated by mild alkaline hydrolysis and added to assay buffer at a concentration 25 times higher than the Ki of the ynenol lactone. Enzyme and substrate were added to the mixture, and neither inhibition nor time-dependent inactivation was observed. Therefore, inactivation was unlikely to occur by enzymatic release of the reactive intermediate followed by nonspecific alkylation outside the active site. 

In enzymatic cleavage from the microspheres either with subtilisin or lipase, the release of the substrate was greater than 60%. Impressively, the release of azo-BSA from the microspheres catalyzed by lipase was greater than 90%. However, nonspecific hydrolysis of azo-BSA by lipase was detected, probably attributed to the similarities between the ester and amide bonds. Also, both reactions were performed in closed systems, contrary to the biological... 

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