title: Purification and characterization of a low molecular mass cysteine proteinase inhibitor from human amniotic fluid.
abstract: We have purified the human low molecular mass cysteine proteinase inhibitor in good yield from amniotic fluid, using ultrafiltration through 100-kDa and 1-kDa cut-off filters, chromatography on Ultrogel AcA 54, and affinity chromatography on alkylated papain-agarose. Approximately 1-4 mg/l of this inhibitor are present in amniotic fluid. The purified inhibitor had an apparent molecular mass of 10.5-12 kDa, as judged by its electrophoretic behavior. Amino acid analysis showed it to be rich in acidic and aliphatic residues and in cysteine. No carbohydrate side-chains could be demonstrated. The purified inhibitor inhibited papain, ficin, cathepsins B, C, and H, the cathepsin B-like enzyme from B16 melanoma cells, and a bovine chromaffin granule enkephalin-converting activity. No inhibition of Ca2-dependent neutral cysteine proteinase, serine- or metallo-proteinases was seen. Analysis of the purified inhibitor by isoelectric focusing revealed 7 major bands with pI values of 7.95, 7.0, 6.7, 6.55, 6.25, 5.5, and 5.2, all of which inhibited papain.
mesh_terms: D000596:Amino Acids / Q000032:analysis; D000653:Amniotic Fluid / Q000032:analysis*; D000818:Animals; D002401:Cathepsin B; D002403:Cathepsins / Q000037:antagonists & inhibitors; D002417:Cattle; D002852:Chromatography, Ion Exchange; D003208:Concanavalin A; D003546:Cysteine Endopeptidases; D015853:Cysteine Proteinase Inhibitors; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D007525:Isoelectric Focusing; D008970:Molecular Weight; D010206:Papain / Q000032:analysis; D011247:Pregnancy; D011480:Protease Inhibitors; D011506:Proteins / Q000032:analysis* / Q000302:isolation & purification
chemical_list: D000596:Amino Acids; D015853:Cysteine Proteinase Inhibitors; D011480:Protease Inhibitors; D011506:Proteins; D003208:Concanavalin A; D002403:Cathepsins; D003546:Cysteine Endopeptidases; D002401:Cathepsin B; D010206:Papain