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Purification and partial characterization of feline pepsinogen

Ursula Tress med vet1, Jörg M. Steiner Dr med vet, PhD2, Craig G. Ruaux BVSc, PhD3, Jan S. Suchodolski Dr med vet4, and David A. Williams VetMB, PhD5
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  • 1 Gastrointestinal Laboratory, Department of Small Animal Medicine and Surgery, College of Veterinary Medicine, Texas A&M University, College Station, TX 77843-4474.
  • | 2 Gastrointestinal Laboratory, Department of Small Animal Medicine and Surgery, College of Veterinary Medicine, Texas A&M University, College Station, TX 77843-4474.
  • | 3 Gastrointestinal Laboratory, Department of Small Animal Medicine and Surgery, College of Veterinary Medicine, Texas A&M University, College Station, TX 77843-4474.
  • | 4 Gastrointestinal Laboratory, Department of Small Animal Medicine and Surgery, College of Veterinary Medicine, Texas A&M University, College Station, TX 77843-4474.
  • | 5 Gastrointestinal Laboratory, Department of Small Animal Medicine and Surgery, College of Veterinary Medicine, Texas A&M University, College Station, TX 77843-4474.

Abstract

Objective—To purify and partially characterize feline pepsinogen (fPG) from the gastric mucosa and compare fPG with PGs of other species.

Sample Population—Stomachs of 6 cats.

Procedure—A crude protein extract was prepared from the gastric mucosa of feline stomachs. Feline PG A was purified by ammonium sulfate precipitation, weak-anion-exchange chromatography, size-exclusion chromatography, and strong-anion exchange chromatography. Partial characterization consisted of estimation of molecular weights (MWs) and isoelectric points, N-terminal amino acid sequencing, and investigation of susceptibility to pepstatin inhibition.

Results—Several fPG A-group isoforms were identified. The MWs of the isoforms ranged from 37,000 to 44,820. Isoelectric points were all < pH 3.0. The proteolytic activity of the activated PGs was inhibited completely by pepstatin in a range of equimolar to 10- fold molar excess. The specific absorbance of fPG A was 1.29. The N-terminal amino acid sequence of the first 25 residues of the predominant fPG A7 had 75%, 72%, 64%, and 56% homology with PG A of dogs, rabbits, cattle, and humans, respectively. Sequences of 4 other fPG A-group isoforms were similar to fPG A7. All isoforms were immunologically cross-reactive with sheep anti-fPG A7 antiserum.

Conclusions and Clinical Relevance—PG A is the only identified type of PG in cats and, similar to pg in other species, comprises multiple isoforms. The availability of fPG A may be used to facilitate the development of an immunoassay to quantify serum fPG A as a potential marker for gastric disorders in cats. (Am J Vet Res 2004;65:1195–1199)

Abstract

Objective—To purify and partially characterize feline pepsinogen (fPG) from the gastric mucosa and compare fPG with PGs of other species.

Sample Population—Stomachs of 6 cats.

Procedure—A crude protein extract was prepared from the gastric mucosa of feline stomachs. Feline PG A was purified by ammonium sulfate precipitation, weak-anion-exchange chromatography, size-exclusion chromatography, and strong-anion exchange chromatography. Partial characterization consisted of estimation of molecular weights (MWs) and isoelectric points, N-terminal amino acid sequencing, and investigation of susceptibility to pepstatin inhibition.

Results—Several fPG A-group isoforms were identified. The MWs of the isoforms ranged from 37,000 to 44,820. Isoelectric points were all < pH 3.0. The proteolytic activity of the activated PGs was inhibited completely by pepstatin in a range of equimolar to 10- fold molar excess. The specific absorbance of fPG A was 1.29. The N-terminal amino acid sequence of the first 25 residues of the predominant fPG A7 had 75%, 72%, 64%, and 56% homology with PG A of dogs, rabbits, cattle, and humans, respectively. Sequences of 4 other fPG A-group isoforms were similar to fPG A7. All isoforms were immunologically cross-reactive with sheep anti-fPG A7 antiserum.

Conclusions and Clinical Relevance—PG A is the only identified type of PG in cats and, similar to pg in other species, comprises multiple isoforms. The availability of fPG A may be used to facilitate the development of an immunoassay to quantify serum fPG A as a potential marker for gastric disorders in cats. (Am J Vet Res 2004;65:1195–1199)