Acid-stable protease inhibiting polypeptides formed from denatured horse plasma by proteolysis.

This research explores how protease inhibiting polypeptides, which are proteins that block enzyme activity, can form from denatured horse plasma through proteolysis, a process that breaks down proteins.

Protease Inhibiting Polypeptides Formation

• Research indicates that the enzymatic digestion of denatured horse plasma with trypsin, an enzyme that breaks down proteins, generated polypeptides. These polypeptides inhibit several enzymes, including trypsin itself, chymotrypsin, and to a lesser extent, kallikrein.
• Two polypeptides were produced through this process, with molecular weights (Mr) of 73,000 and 24,000.
• The number, enzyme specificity, and Mr of these inhibitory polypeptides found in horses differed from those known for humans.

Purification of Polypeptides

• The inhibitory polypeptides were purified using two methods: affinity chromatography on Sepharose-trypsin, a technique that separates biological molecules based on their interaction with specific ligands, and gel filtration via Sephadex G-75, a technique that separates molecules based on their size.

Generation of Polypeptides Using Various Enzymes

• The research also discovered that protease inhibitory polypeptides can be generated similarly using various enzymes including chymotrypsin, elastase, proteinase K, pronase, collagenase, papain, and subtilisin.
• The number and the electrophoretic migration, a technique used to separate and characterize molecules based on their rate of movement in an electric field, of these inhibitory polypeptides varied when different enzymes were used.

Specificity and Immunological Relation

• The activities of these polypeptides were specific; they only inhibited trypsin, chymotrypsin and marginally kallikrein, irrespective of the enzyme used for their generation.
• Immunologically, none of these polypeptides were found to be related to native plasma proteins or plasma protease inhibitors, indicating that they may be a unique type of protein.