Enzyme antigens associated with pigeon breeder's disease. II. Isolation and characterization of acidic hydrolases

Daniel J. McCormick, Walter W. Fredricks, Thomas H. Tebo, Nickolas J. Calvanico

Research output: Contribution to journalArticlepeer-review

Abstract

The hydrolytic enzymes in pigeon dropping extracts (PDE) were separated into basic and acidic components by DEAE-cellulose chromatography. Six distinct hydrolytic activities were isolated from the acidic group of enzymes. Elastase, trypsin, and two forms of collagenase were the proteases identified. An esterase and a phospholipase were also detected. The sera of symptomatic pigeon breeders, analyzed by crossed immunoelectrofocusing techniques, were shown to contain antibodies to the enzymes trypsin, collagenase, and esterase in the acidic fractions of PDE by staining the immune precipitates with specific chromogenic substrates. These enzymes were purified by preparative isoelectric focusing, affinity chromatography, and gel filtration. The purified elastase hydrolyzed elastin and consisted of a single polypeptide chain (Mr=23,100). Trypsin hydrolyzed a synthetic arginine substrate, but not elastin or collagen. Its size (Mr=27,500) and subunit structure were similar to those reported here for elastase. Both enzymes were inhibited by α-1-antitrypsin, Nα-p-tosyl-L-lysine chloromethylketone, and phenylmethylsulfonyl fluoride. Two distinct collagenases were found; both cleaved bovine collagen. The high-molecular-weight (HMW) collagenase was a glycoprotein (Mr=117,500) and consisted of three polypeptide chains (Mr=37,100); a low-molecular-weight (LMW) collagenase (Mr=12,500) was also isolated. The HMW collagenase was inhibited by EDTA, p-chloromercuribenzoic acid, and phenylmethylsulfonyl fluoride, but not by α-1-antitrypsin. The source of these enzymes as well as their relationship to the basic hydrolytic enzymes of PDE are discussed.

Original languageEnglish (US)
Pages (from-to)309-325
Number of pages17
JournalJournal of Protein Chemistry
Volume3
Issue number3
DOIs
StatePublished - Jun 1 1984

Keywords

  • allergic alveolitis
  • hypersensitivity pneumonitis
  • pulmonary allergens

ASJC Scopus subject areas

  • Biochemistry

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