Plasma membrane-bound and lysosomal peptidases in human alveolar macrophages.
Alveolar macrophages protect the lungs against noxious agents. Proteases and peptidases are essential for this defense and many metabolic activities. Human alveolar macrophages were evaluated for the presence of six important peptidases. Deamidase, a serine peptidase identical with the lysosomal protective protein and possibly with cathepsin A, had high specific activity in alveolar macrophages and is also present in cultured mouse J774A.1 and human U937 cells, used for the sake of comparison. In fractionated J774A cells, most of the deamidase activity was in the lysosomal fraction and in the final supernatant. Deamidase in human alveolar macrophages, obtained by bronchoalveolar lavage from 23 patients, cleaved dansyl-Phe-Leu-Arg at a rate of 2.26 mumol/h/mg protein and hydrolyzed the chemotactic peptide N-f-Met-Leu-Phe even faster, at a rate of 53.1 mumol/h/mg protein, the highest activity for this enzyme with any of the cells we tested. Rabbit antiserum, elicited with the recombinant partial sequence of the enzyme, immunoprecipitated 77-88% of the macrophage deamidase. In immunocytochemistry, this antiserum localized deamidase within the human macrophages. The enzyme was inhibited by diisopropylfluorophosphate (DFP; 1 mM) and by ebelactone B (10 microM), noncompetitively. The mRNA of deamidase was detected in mouse macrophages by Northern blot; the two protein chains of deamidase were shown in human macrophages by Western blot. In addition, two other serine peptidases were also highly active in macrophages: dipeptidyl peptidase IV (1.38 mumol/h/mg protein) and prolylcarboxypeptidase (0.72 mumol/h/mg protein). The activity of plasma membrane zinc metallopeptidases, neutral endopeptidase 24.11 and carboxypeptidase M, in contrast, was low or absent (angiotensin I converting enzyme; kininase II).(ABSTRACT TRUNCATED AT 250 WORDS)
Jackman
H L
HL
Department of Pharmacology, University of Illinois College of Medicine, Chicago 60612, USA.
Tan
F
F
Schraufnagel
D
D
Dragović
T
T
Dezsö
B
B
Becker
R P
RP
Erdös
E G
EG
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002268:Carboxypeptidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D043402:Cathepsin A; D002478:Cells, Cultured / Q000201:enzymology; D002630:Chemotactic Factors / Q000378:metabolism; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D007150:Immunohistochemistry; D008247:Lysosomes / Q000201:enzymology*; D016676:Macrophages, Alveolar / Q000201:enzymology*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism; D011233:Precipitin Tests; D013379:Substrate Specificity
D002630:Chemotactic Factors; D008565:Membrane Proteins; D010455:Peptides; D002268:Carboxypeptidases; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; C468629:CTSA protein, human; D043402:Cathepsin A