Iselin, Beat M. and Niemann, Carl (1950) A procedure for the determination of proteolytic activity. Journal of Biological Chemistry, 182 (2). pp. 821-828. ISSN 0021-9258. http://resolver.caltech.edu/CaltechAUTHORS:ISEjbc50a
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The difficulties introduced by the desire to maintain a constant pH during an enzyme-catalyzed hydrolysis of peptide-like substrates and at the same time to determine the extent of hydrolysis by an acid-base titration have been pointed out (1), but to date no completely satisfactory solution of the problem has been given. With those enzymes whose pH optima lie in the region between pH 7.5 to 8.5, e.g. trypsin and chymotrypsin, the poor buffering capacity of phosphate in this region prompted us, as it has others (2-5), to consider the use of organic amines whose pK’alpha values were near to or identical with the pH optimum of the enzyme being used. In the course of such studies it soon became evident that coincidental use of a suitable primary or secondary amine buffer system and a formol titration (1) would insure adequate buffering capacity with low buffer concentration during the hydrolysis and at the same time permit the final acid-base titration to be conducted under nearly ideal conditions. In this communication we shall limit the discussion to results obtained with chymotrypsin and specific acylated-a-amino acid amide substrates, since the application of the general method to other proteolytic enzymes and other types of substrates will be obvious.
|Additional Information:||Copyright © 1950 by the American Society of Biological Chemists (Received for publication, September 27, 1949) Supported in part by a grant from Eli Lilly and Company. Contribution No. 1334 from the Gates and Crellin Laboratories of Chemistry, California Institute of Technology, Pasadena|
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|Deposited On:||29 Aug 2006|
|Last Modified:||26 Dec 2012 09:00|
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