Isolation and characterization of alpha 1-protease inhibitor from canine plasma.

Abstract

OBJECTIVE:

To improve a previously described purification process by producing a higher yield and purity of alpha 1-protease inhibitor (alpha 1-PI) from canine plasma.

ANIMALS:

Plasma pool from 10 clinically normal male dogs.

PROCEDURE:

Canine alpha 1-PI was purified by use of ammonium sulfate precipitation, ion-exchange chromatography, and 3 affinity chromatographic procedures: concanavalin A-Sepharose, thiol, and hemoglobin-Sepharose. Characterization was performed by gel electrophoresis, isoelectric focusing, and immunoblot analysis. The N-terminal amino acid sequence was obtained by use of the Edman degradation method and a gas amino acid sequencer.

RESULTS:

Canine alpha 1-PI was purified with a yield of approximately 7% and a 54-fold increase in specific inhibitory activity. The inhibitor had a molecular weight of 59,000 and had 2 major patterns after isoelectric focusing: fast and intermediate in homozygous and/or heterozygous forms. Edman degradation revealed glutamic acid as the starting amino acid from the N-terminal sequence. Homologies of the N-terminal sequence of canine alpha 1-PI with those of sheep, horse, and human alpha 1-protease inhibitors were 54, 46, and 41%, respectively.

CONCLUSIONS:

Canine protease inhibitor is analogous to the alpha 1-protease inhibitors of sheep, human beings, and mice in terms of molecular weight, amino acid composition, and inhibitory activity against trypsin. Although the method described had a yield of 7%, the final product retained inhibitory activity and was pure.

CLINICAL RELEVANCE:

The availability of pure canine alpha 1-PI, as well as the specific antibodies, will facilitate studies on the fecal excretion and structural heterogeneity of this protein in dogs with naturally acquired protein-losing enteropathy.

• Authors: T Melgarejo, D A Williams, G Griffith
• Journal: American journal of veterinary research (Am J Vet Res) Vol. 57 Issue 3 Pg. 258-63 (Mar 1996) ISSN: 0002-9645 [Print] United States
• PMID: 8669751 (Publication Type: Comparative Study, Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antibodies
• alpha 1-Antitrypsin

Topics

• Amino Acid Sequence
• Animals
• Antibodies
• Chromatography, Affinity
• Chromatography, Ion Exchange
• Dogs
• Electrophoresis, Polyacrylamide Gel
• Heterozygote
• Homozygote
• Horses
• Humans
• Immunoblotting
• Isoelectric Focusing
• Male
• Mice
• Molecular Sequence Data
• Sequence Homology, Amino Acid
• alpha 1-Antitrypsin (chemistry, genetics, isolation & purification)