Aminopeptidases from human leucocytes.
Six aminopeptidases differing in enzymic specificity against various L-amino acid-4-nitroanilides were detected and isolated from the cytosol of leucocytes collected from the buffy coat of human blood. The different enzymes were separated by one step of chromatography on DEAE-Sephacel and were further purified by gel filtration on Sephacryl S-300. The main aminopeptidases of the cytosol were designated aminopeptidases 1, 2, 4 and 5 (AP 1, AP 2, AP 4, AP 5) on the basis of their elution sequence from the first ion-exchange chromatography column on DEAE-Sephacel. Aminopeptidase 1 appeared to be a strongly sulfhydryl-dependent leucine aminopeptidase, activatable by thiol reagents. The enzyme was inhibited by p-chloromercuribenzoate. Its molecular mass was estimated to be 150 kDa. Aminopeptidase 2 showed high specificity for proline-4-nitroanilide. This enzyme was inhibited by p-chloro-mercuribenzoate and bestatin. It exhibited a molecular mass of 70 kDa. Aminopeptidase 4 designated the activities of two different enzymes of apparent molecular masses of 220 and 70 kDa which could be further separated by gel filtration. Aminopeptidase 5 exhibited the properties alike aminopeptidase B with high specific hydrolytic activity against the 4-nitroanilides of lysine and arginine. The molecular mass was estimated to be 90 kDa. Aminopeptidase 3 was a minor component in the cytosol and could be identified as an extracellular leucocyte plasma membrane constituent. The enzyme exhibited properties of a metallo proteinase and could be inhibited by EDTA. The molecular mass was estimated to be 250 kDa.
D000626:Aminopeptidases / Q000097:blood* / Q000302:isolation & purification; D001798:Blood Proteins / Q000032:analysis; D002850:Chromatography, Gel / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D003600:Cytosol / Q000201:enzymology; D006801:Humans; D007527:Isoenzymes / Q000097:blood / Q000302:isolation & purification; D007962:Leukocytes / Q000201:enzymology*; D008970:Molecular Weight; D013056:Spectrophotometry, Ultraviolet
D001798:Blood Proteins; D007527:Isoenzymes; D000626:Aminopeptidases