Leishmania: Immunochemical Comparison of the Secretory (Extracellular) Acid Phosphatases from Various Species

Abstract

Promastigotes of all Leishmania species, except most Leishmania major stocks, synthesize and secrete extracellular soluble acid phosphatases [SAcP(s)] which are capable of dephosphorylating a wide range of substrates and, consequently, of modifying the parasite environment. To assess their antigenic/structural relatedness, the SAcP(s) from various species were compared immunochemically using both a monospecific rabbit antiserum and several monoclonal antibodies made against the purified promastigote SAcP of a cloned line of Leishmania donovani donovani. Results obtained from antibody-bridged enzyme activity assays demonstrated that these reagents quantitatively immunoprecipitated the enzymatic activities of SAcP(s) from 18 different W.H.O. reference stocks of Leishmania, including two L. major strains. Those results showed, for the first time, that all SAcP(s) possessed some common cross-reactive antigenic epitopes. Immunoprecipitates obtained from [35S]methionine, metabolically labeled promastigote culture supernatants of the various species, were analyzed by SDS-PAGE/fluorography. These analyses showed that marked differences existed among the SAcP(s) both in their relative mobilities and the number of constituent bands resolved. Tunicamycin treatment resulted in a significant reduction in the apparent molecular weights of SAcP(s) from three different isolates, confirming the presence of N-linked carbohydrate side chains in these enzymes. In addition, the SAcP released by L. donovani intracellular amastigotes was also immunoprecipitated with the monospecific rabbit antisera from the culture supernatant of infected macrophages. Cumulatively, these data demonstrate that despite individual species variations, the SAcP(s) from all Leishmania tested have retained certain common antigenic/structural epitopes and functional protein domains. Such conservation among SAcP(s) suggests that this enzyme must play an essential role in the survival of promastigotes and amastigotes of all Leishmania species.