Heterogeneity of cysteine proteinases in Leishmania braziliensis and Leishmania major.
Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with acrylamide copolymerized with gelatin (substrate-SDS-PAGE) was used to compare promastigote proteinases from two Leishmania species (L. braziliensis and L. major-like). Substrate-SDS-PAGE resolved at least 6 distinct proteinase activities with relative molecular masses between 20 and 65 kDa. The profile of proteinase activity was the same for both species, although qualitative differences were observed. L. major-like expressed a low molecular weight (20 kDa) proteinase with maximum activity at pH 7.0, which was demonstrable from pH 5.0 to pH 8.0. The 20-kDa protease was recovered in the detergent phase of TX-114 and was inhibited by the sulfhydryl group-blocking reagent E-64 (2.5 mM) and the non-specific inhibitor iodoacetic acid (1 mM). Pepstatin (1 microM) and PMSF (2.5 mM) did not inhibit the 20-kDa enzyme. The present study suggests that both Leishmania species studied express hydrophobic cysteine proteases of different molecular weights, since about 2 x 10(7) parasites were analyzed in each lane and the proteolytic activity developed at 37 degrees C for 16 h.
Alves
C R
CR
Departamento de Ciências Biológicas, Escola Nacional de Saúde Pública, Rio de Janeiro, Brasil.
Marzochi
M C
MC
Giovanni-de-Simone
S
S
D000818:Animals; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D007892:Leishmania braziliensis / Q000201:enzymology*; D007895:Leishmania tropica / Q000201:enzymology*; D008970:Molecular Weight
D003546:Cysteine Endopeptidases