Thermolysin proteolysis

Proteolysis of peptides and proteins by enzymes occurs in a selective or nonselective manner. Chymotrypsin, trypsin, lysyl endopeptidase, Staphylococcus aureus V8 protease, and en-dopeptidase Asp-N are frequently listed as selective enzymes, whereas thermolysin, pepsin, subtilisin, and elastase belong to the nonselective enzymes, although thermolysin preferentially cleaves peptide bonds before hydrophobic residues. 

Limited proteolysis by thermolysin, papain, subtilsin and a-chymotrypsin of the plasma-membrane-bound adenylate cyclase from rat liver had no effect on activity when Mn2+ was the cofactor, but stimulated activity in the presence of Mg2+. However, the addition of Mg2+ over MnATP did not result in stimulation on proteolysis. The simplest interpretation is that proteolysis exerts its main effect on the regulatory components of adenylate cyclase.329 Adenylate cyclase, solubilized from rat liver membranes by Lubrol PX or deoxycholate, loses its regulatory components, and becomes much more specific for MnATP.330... 

Limited proteolysis by thermolysin, papain, subtilsin and a-chymotrypsin of the plasma-membrane-bound adenylate cyclase from rat liver had no effect on activity w hen Mn was the... 

Table 6. ACE inhibition by plant peptides and peptides derived from plant protein proteolysis Peptide sequences are given in both the three-letter and one-letter codes. Triticum aestivum (wheatgerm) protein was proteolysed by Bacilllus lichenoformis alkaline protease [i99] and Zea mays (com seed) a-zein was proteolysed by thermolysin [203, 204],...

Matthews et al. have determined the structure of the extracellular proteolytic enzyme, thermolysin, to 2.3 A resolution. This enzyme of molecular weight 37 500 contains one zinc and four calcium ions, and is interesting because of its imusual heat stability. The active site contains a zinc atom tetrahedrally co-ordinated to His-142, His-146, Glu-166, and a water molecule, and in this way resembles carboxypeptidase A. The precise details of the co-ordination of the calcium ions are not reported. However, it is of interest that two have a centre-to-centre distance of 3.8 A and are located within an interior region of the protein, surrounded by carbonyl and carboxylate groups. Loss of calcium does not hinder proteolysis at room temperature, but the enzyme is no longer heat stable. Preliminary X-ray data are also reported for the acidic protease from the fungus Rhizopus chinensis. ... 

Two non-inhibitory proteinase—proteinase inhibitor combinations exhibit destabilization of inhibitor proteins in the DSC (Zahnley, unpublished). Ovoinhibitor was rapidly degraded by thermolysin, as indicated by decreases in both T(j and area (AHd) of its endotherm. Chicken ovomucoid was slowly degraded by proteinase K (Fig. 9), a fungal serine proteinase that attacks many native proteins. Proteinase K, in accord with its subtilisin-1ike specificity (Kraus and Femfert, 1976), was inhibited by chicken ovoinhibitor, but not by chicken ovomucoid. In these non-inhibited systems, proteolysis can destabilize the inhibitor (substrate) protein, provided the susceptible bonds are accessible to the proteinase. Relevance of these examples to food systems derives from the role of proteolysis in protein deterioration (Feeney, 1980) and digestion. 

After reaction with the labeled reagents, the protein is treated with proteolytic enzymes thermolysin, chymotrypsin, and trypsin have been used for proteolysis of the labeled macromolecule. The resulting peptides are separated by high-pressure liquid chromatography (HPLC), and the amount of radioactivity of each eluted peak is measured. Then the labeled peptides are analyzed after acid hydrolysis and the amount of radioactive label incorporated into individual amino acid side chains is measured. 

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