TY - JOUR
T1 - Catalytic properties of cysteine proteinases from Trypanosoma cruzi and Leishmania infantum
T2 - A pre-steady-state and steady-state study
AU - Ascenzi, Paolo
AU - Bocedi, Alessio
AU - Visca, Paolo
AU - Antonini, Giovanni
AU - Gradoni, Luigi
PY - 2003/9/26
Y1 - 2003/9/26
N2 - Cysteine proteinases are relevant to several aspects of the parasite life cycle and of parasite-host relationship. Moreover, they appear as promising targets for antiparasite chemotherapy. Here, the first quantitative investigation on the steady-state and pre-steady-state kinetics of the papain-like cysteine proteinases from epimastigotes of Trypanosoma cruzi (cruzipain), the agent of Chagas' disease, and from promastigotes of Leishmania infantum, an agent of visceral and cutaneous leishmaniases, is reported. The results indicate that kinetics for the parasite proteinase catalyzed hydrolysis of N-α -benzyloxycarbonyl-L-phenylalanyl-L-arginine-(7-amino-4-methylcoumarin) may be consistently fitted to the minimum three-step mechanism involving the acyl·enzyme intermediate E·P: KS k+2 k +3 E+S ↔ E:S → E·P → E+P2 + P 1 At neutral pH, the k+3 step (deacylation process) is rate limiting in enzyme catalysis, whereas, at pH+2 step (acylation process) becomes rate limiting. This illustrates the potential danger in interpreting both kcat versus pH profile, given that the acylation or the deacylation step is rate limiting throughout the whole pH range explored, and Km as the true affinity constant for the E:S complex formation. Comparison with the steady-state and pre-steady-state kinetics of homologous plant enzymes suggests that the parasite cysteine proteinase catalytic behavior appears to be of general significance.
AB - Cysteine proteinases are relevant to several aspects of the parasite life cycle and of parasite-host relationship. Moreover, they appear as promising targets for antiparasite chemotherapy. Here, the first quantitative investigation on the steady-state and pre-steady-state kinetics of the papain-like cysteine proteinases from epimastigotes of Trypanosoma cruzi (cruzipain), the agent of Chagas' disease, and from promastigotes of Leishmania infantum, an agent of visceral and cutaneous leishmaniases, is reported. The results indicate that kinetics for the parasite proteinase catalyzed hydrolysis of N-α -benzyloxycarbonyl-L-phenylalanyl-L-arginine-(7-amino-4-methylcoumarin) may be consistently fitted to the minimum three-step mechanism involving the acyl·enzyme intermediate E·P: KS k+2 k +3 E+S ↔ E:S → E·P → E+P2 + P 1 At neutral pH, the k+3 step (deacylation process) is rate limiting in enzyme catalysis, whereas, at pH+2 step (acylation process) becomes rate limiting. This illustrates the potential danger in interpreting both kcat versus pH profile, given that the acylation or the deacylation step is rate limiting throughout the whole pH range explored, and Km as the true affinity constant for the E:S complex formation. Comparison with the steady-state and pre-steady-state kinetics of homologous plant enzymes suggests that the parasite cysteine proteinase catalytic behavior appears to be of general significance.
KW - Leishmania infantum
KW - Parasite cysteine proteinase
KW - Pre-steady-state kinetics
KW - Steady-state kinetics
KW - Trypanosoma cruzi
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U2 - 10.1016/j.bbrc.2003.08.015
DO - 10.1016/j.bbrc.2003.08.015
M3 - Article
C2 - 12963041
AN - SCOPUS:0041328861
VL - 309
SP - 659
EP - 665
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
SN - 0006-291X
IS - 3
ER -