Native and unfolded forms of ribonuclease, lysozyme, and trypsin have been subjected to partial proteolysis with trypsin or pronase. Soluble peptides are formed from the mixed disulfides of lysozyme or ribonuclease, with cystine at a rate intermediate between the rates of soluble peptide formation from native and other unfolded forms of these two enzymes. Active and inactive products isolated from irradiated lysozyme samples have been subjected to limited proteolysis employing trypsin and to extensive proteolysis employing combined peptic-tryptic hydrolysis. Alterations in conformation or motility of peptide chains have been detected in all products tested, including the one most nearly corresponding chromatographically to native lysozyme. Detection of alterations is dependent upon following more than one parameter of proteolysis. A change in the rate or extent of peptide bond cleavage is not always accompanied by a change in rate of proteolytic release of trichloroacetic acid soluble peptides. Of the three proteases studied, the use of trypsin most consistently reveals differences between the protease substrates. The use of trypsin is indicated for the production of a limited number of large peptide intermediates from lysozyme derivatives for isolation and characterization to reveal small conformational changes. The use of a combination of pepsin and trypsin is indicated for the production of a mixture of small peptides containing disulfide bonds rearranged in lysozyme derivatives from their position in native lysozyme.

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