Resistance of horse alpha 1-proteinase inhibitor to perchloric acid denaturation and a simplified purification procedure resulting therefrom.

This study discovers that horse alpha 1-proteinase inhibitor is unexpectedly resistant to denaturation (unfolding of proteins) caused by perchloric acid. This surprising characteristic made it possible for the researchers to develop a simplified procedure for purifying this inhibitor, and the resulting preparations were observed to be immunogenically pure.

The Resistance to Perchloric Acid

• The paper discusses a unique characteristic of the horse alpha 1-proteinase inhibitor. Despite the addition of perchloric acid, the inhibitor did not precipitate or become inactive. Precipitation usually occurs when proteins unfold or denature, but such a phenomenon was not seen here.
• Interestingly, even the isoinhibitors, which are different forms of the alpha 1-proteinase inhibitor, were not changed by the perchloric acid. Typically, a change in protein structure would be expected.

Simplified Purification Procedure

• The resistance observed led researchers to develop a novel, simpler process to purify horse alpha 1-proteinase inhibitor. This process involved removal of the bulk of plasma proteins by perchloric acid precipitation and gel filtrations.
• Gel filtration on Sephadex G-75 and G-200 was used as part of the purification process. Gel filtration is a type of chromatography which separates proteins based on their size. Sephadex is a type of gel used in this method.

Results of Purification

• The preparations of alpha 1-proteinase inhibitor that resulted from this new purification method were found to be immunogenically pure. This means they were able to invoke an immune response, which implies they maintained their functionality.
• The simple purification procedure enabled the comparison of isoinhibitors of alpha 1-proteinase inhibitor, demonstrating that they were immunologically identical. This suggests that different forms of the inhibitor are indistinguishable in terms of their immune system interactions.