| Full text | |
| Author(s): |
Gomes, Mayrim M.
[1, 2]
;
Budu, Alexandre
[3]
;
Ventura, Priscilla D. S.
[1]
;
Bagnaresi, Piero
[3]
;
Cotrin, Simone S.
[3]
;
Cunha, Rodrigo L. O. R.
[4]
;
Carmona, Adriana K.
[3]
;
Juliano, Luiz
[3]
;
Gazarini, Marcos L.
[1]
Total Authors: 9
|
| Affiliation: | [1] Univ Fed Sao Paulo, Dept Biociencias, Santos, SP - Brazil
[2] Univ Fed Sao Paulo, Escola Paulista Med, Dept Microbiol Immunol & Parasitol, Sao Paulo - Brazil
[3] Univ Fed Sao Paulo, Dept Biofis, Sao Paulo - Brazil
[4] Univ Fed ABC, Ctr Ciencias Nat & Humanas, Santo Andre, SP - Brazil
Total Affiliations: 4
|
| Document type: | Journal article |
| Source: | Analytical Biochemistry; v. 468, p. 22-27, JAN 1 2015. |
| Web of Science Citations: | 5 |
| Abstract | |
In the intraerythrocytic trophozoite stages of Plasmodium falciparum, the calcium-dependent cysteine protease calpain (Pf-calpain) has an important role in the parasite calcium modulation and cell development. We established specific conditions to follow by confocal microscopy and spectrofluorimetry measurements the intracellular activity of Pf-calpain in live cells. The catalytic activity was measured using the fluorogenic Z-Phe-Arg-MCA (where Z is carbobenzoxy and MCA is 4-methylcoumaryl-7-amide). The calmodulin inhibitor calmidazolium and the sarcoplasmic reticulum calcium ATPase inhibitor thapsigargin were used for modifications in the cytosolic calcium concentrations that persisted in the absence of extracellular calcium. The observed calcium-dependent peptidase activity was greatly inhibited by specific cysteine protease inhibitor E-64 and by the selective calpain inhibitor ALLN (N-acetyl-t-leucyl-L-leucyl-L-norleucinal). Taken together, we observed that intracellular Pf-calpain can be selectively detected and is the main calcium-dependent protease in the intraerythrocytic stages of the parasite. The method described here can be helpful in cell metabolism studies and antimalarial drug screening. (C) 2014 Elsevier Inc. All rights reserved. (AU) | |
| FAPESP's process: | 10/09264-2 - Participation of phosphorylation and calcium homeostasis in intracellular proteolytic activity modulation in Plasmodium chabaudi |
| Grantee: | Mayrim Machado Gomes Smaul |
| Support Opportunities: | Scholarships in Brazil - Scientific Initiation |
| FAPESP's process: | 11/15287-8 - Proteolytic activity study in Plasmodium and modulation of kallikrein-kinin system in infected livers |
| Grantee: | Mayrim Machado Gomes Smaul |
| Support Opportunities: | Scholarships in Brazil - Master |
| FAPESP's process: | 13/12913-0 - Calcium-calmodulin of Plasmodium falciparum: identification of ligands and participation in the host-parasite signalling. |
| Grantee: | Alexandre Budu |
| Support Opportunities: | Scholarships in Brazil - Post-Doctoral |
| FAPESP's process: | 09/53840-0 - Fluorescence system for investigations of physiological and pathophysiological aspects in cellular models |
| Grantee: | Adriana Karaoglanovic Carmona |
| Support Opportunities: | Multi-user Equipment Program |