CD13 is a membrane-bound cell surface glycoprotein that was originally recognized as a marker for subsets of normal and malignant myeloid cells, but later found on other cells and tissues. Sequence comparisons of the cloned cDNA have shown that CD13 is identical to aminopeptidase N (APN; EC 3.4.11.2), a type II membrane-spanning protein belonging to the M1 family of zinc-metallopeptidase which cleaves N-terminal neutral and basic amino acids from peptides and proteins. This ectoenzyme can cleave bioactive proteins on the cell surface, including cytokines, to either activate or inactivate them, and it has been involved in down-regulation of signal peptides, angiogenesis, inflammation, cell signalling and even as a receptor for certain viruses (e.g. coronavirus 229E) enabling host cell infection. Soluble aminopeptidase activity found in serum appears to procede from native membranous APN which is cleaved off by limited proteolysis and is characterized by the absence of the hydrophobic N-terminal polypeptide chain, responsible for anchoring the protein to the membrane.
Since antimalarial drugs are used to treat milder cases of rheumatoid arthritis and systemic lupus erythematosus due to their antiinflammatory properties and since aminopeptidases seem to be involved in the terminal stages of globin degradation in Plasmodium falciparum we have investigated the effects of these substances on aminopeptidase activity of human serum in a fluorometric assay that use amino acyl β-naphthylamide (βNA) substrates.
Serum aminopeptidase activities from human healthy donnors varied depending on the substrate used (alanine-β-naphthylamide, arginine-β-naphthylamide or leucine-β-naphthylamide), ala-βNA being the substrate more actively hydrolysed (17.7 ± 1.0 nm min-1 ml-1; mean ± S.E.M.); arg-βNA (5.4 ± 0.2 nm min-1 ml-1) and leu-βNA (5.9 ± 0.6 nm min-1 ml-1) were hydrolysed to a lesser degree.
Quinine (hydrochloride) and chloroquine (diphosphate salt) inhibited AP activities in a dose-related manner independently of the substrate used. In general, chloroquine (IC50: 71.7 mM; substrate: ala-βNA) was more effective inhibiting AP activity than quinine (IC50: 149.6 mM; substrate: ala-βNA). Nevertheless, the lowest IC50 was obtained when the substrate was arg-βNA. In all the cases data can be adapted to the negative exponential curve y = a0 + a1e (-x/a2) with a coefficient of determination > 0.99 in every case.
A wide variety of mechanisms of antiinflammatory action have been proposed for antimalarial agents. Reduced T lymphocyte mitogen responsiveness, reduced leucocyte chemotaxis, stabilization of lysosomal membranes or trapping free radicals are the most commonly proposed. The inhibition of aminopeptidase activities by these agents suggests that these enzymes may play a role in the mechanism by which antimalarial drugs have therapeutic effects.
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