Trypanosoma cruzi: Partial characterization of minor cruzipain isoforms non-adsorbed to Concanavalin A-Sepharose
The present paper reports the partial characterization of a subset of atypical cruzipain molecules which do not bind to Concanavalin A-Sepharose column. They are present in different strains of epimastigote forms of Trypanosoma cruzi and represent a 2-4% of total cruzipain. They were purified by aff...
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todo:paper_00144894_v104_n3-4_p122_Duschak2023-10-03T14:12:46Z Trypanosoma cruzi: Partial characterization of minor cruzipain isoforms non-adsorbed to Concanavalin A-Sepharose Duschak, V.G. Barboza, M. Couto, A.S. 1,10 phenanthroline aprotinin azo compound bovine serum albumin casein chromogenic substrate concanavalin A cruzipain cystatin cysteine proteinase edetic acid fluoride gelatin hemoglobin isoenzyme lectin leupeptin n [n (3 carboxyoxirane 2 carbonyl)leucyl]agmatine oligosaccharide pepstatin peptide derivative phenylmethylsulfonylfluoride polyclonal antibody sepharose tosyllysyl chloromethyl ketone unclassified drug acidity adsorption affinity chromatography alkalinity anion exchange chromatography article controlled study enzyme activity enzyme analysis enzyme assay enzyme purification enzyme substrate epimastigote hydrolysis nonhuman priority journal protein protein interaction Trypanosoma cruzi Bovinae Trypanosoma Trypanosoma cruzi The present paper reports the partial characterization of a subset of atypical cruzipain molecules which do not bind to Concanavalin A-Sepharose column. They are present in different strains of epimastigote forms of Trypanosoma cruzi and represent a 2-4% of total cruzipain. They were purified by affinity chromatography on Cystatin-Sepharose, recognized by the polyclonal anti-cruzipain serum, and their activity in gelatin-containing gels was completely abolished by E-64, TLCK, leupeptin, and aprotinin but not by PMSF, pepstatin A, EDTA or 1,10-phenantroline. These cysteine proteinases, as well as cruzipain showed to be endoproteinases able to hydrolize azocasein, hemoglobin, and bovine serum albumin at acidic pHs. However, evidences are presented indicating that this subset of cruzipain isoforms were also able to use the same blocked chromogenic peptidyl substrates than cruzipain at similar optimal alkaline pH values although with a different order of preference. Moreover, they showed a different oligosaccharide pattern after enzymatic treatment by high pH anion exchange chromatography, suggesting that this structural difference may account for the atypical behaviour in the lectin columm. © 2003 Elsevier Inc. All rights reserved. JOUR info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_00144894_v104_n3-4_p122_Duschak |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
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Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
1,10 phenanthroline aprotinin azo compound bovine serum albumin casein chromogenic substrate concanavalin A cruzipain cystatin cysteine proteinase edetic acid fluoride gelatin hemoglobin isoenzyme lectin leupeptin n [n (3 carboxyoxirane 2 carbonyl)leucyl]agmatine oligosaccharide pepstatin peptide derivative phenylmethylsulfonylfluoride polyclonal antibody sepharose tosyllysyl chloromethyl ketone unclassified drug acidity adsorption affinity chromatography alkalinity anion exchange chromatography article controlled study enzyme activity enzyme analysis enzyme assay enzyme purification enzyme substrate epimastigote hydrolysis nonhuman priority journal protein protein interaction Trypanosoma cruzi Bovinae Trypanosoma Trypanosoma cruzi |
| spellingShingle |
1,10 phenanthroline aprotinin azo compound bovine serum albumin casein chromogenic substrate concanavalin A cruzipain cystatin cysteine proteinase edetic acid fluoride gelatin hemoglobin isoenzyme lectin leupeptin n [n (3 carboxyoxirane 2 carbonyl)leucyl]agmatine oligosaccharide pepstatin peptide derivative phenylmethylsulfonylfluoride polyclonal antibody sepharose tosyllysyl chloromethyl ketone unclassified drug acidity adsorption affinity chromatography alkalinity anion exchange chromatography article controlled study enzyme activity enzyme analysis enzyme assay enzyme purification enzyme substrate epimastigote hydrolysis nonhuman priority journal protein protein interaction Trypanosoma cruzi Bovinae Trypanosoma Trypanosoma cruzi Duschak, V.G. Barboza, M. Couto, A.S. Trypanosoma cruzi: Partial characterization of minor cruzipain isoforms non-adsorbed to Concanavalin A-Sepharose |
| topic_facet |
1,10 phenanthroline aprotinin azo compound bovine serum albumin casein chromogenic substrate concanavalin A cruzipain cystatin cysteine proteinase edetic acid fluoride gelatin hemoglobin isoenzyme lectin leupeptin n [n (3 carboxyoxirane 2 carbonyl)leucyl]agmatine oligosaccharide pepstatin peptide derivative phenylmethylsulfonylfluoride polyclonal antibody sepharose tosyllysyl chloromethyl ketone unclassified drug acidity adsorption affinity chromatography alkalinity anion exchange chromatography article controlled study enzyme activity enzyme analysis enzyme assay enzyme purification enzyme substrate epimastigote hydrolysis nonhuman priority journal protein protein interaction Trypanosoma cruzi Bovinae Trypanosoma Trypanosoma cruzi |
| description |
The present paper reports the partial characterization of a subset of atypical cruzipain molecules which do not bind to Concanavalin A-Sepharose column. They are present in different strains of epimastigote forms of Trypanosoma cruzi and represent a 2-4% of total cruzipain. They were purified by affinity chromatography on Cystatin-Sepharose, recognized by the polyclonal anti-cruzipain serum, and their activity in gelatin-containing gels was completely abolished by E-64, TLCK, leupeptin, and aprotinin but not by PMSF, pepstatin A, EDTA or 1,10-phenantroline. These cysteine proteinases, as well as cruzipain showed to be endoproteinases able to hydrolize azocasein, hemoglobin, and bovine serum albumin at acidic pHs. However, evidences are presented indicating that this subset of cruzipain isoforms were also able to use the same blocked chromogenic peptidyl substrates than cruzipain at similar optimal alkaline pH values although with a different order of preference. Moreover, they showed a different oligosaccharide pattern after enzymatic treatment by high pH anion exchange chromatography, suggesting that this structural difference may account for the atypical behaviour in the lectin columm. © 2003 Elsevier Inc. All rights reserved. |
| format |
JOUR |
| author |
Duschak, V.G. Barboza, M. Couto, A.S. |
| author_facet |
Duschak, V.G. Barboza, M. Couto, A.S. |
| author_sort |
Duschak, V.G. |
| title |
Trypanosoma cruzi: Partial characterization of minor cruzipain isoforms non-adsorbed to Concanavalin A-Sepharose |
| title_short |
Trypanosoma cruzi: Partial characterization of minor cruzipain isoforms non-adsorbed to Concanavalin A-Sepharose |
| title_full |
Trypanosoma cruzi: Partial characterization of minor cruzipain isoforms non-adsorbed to Concanavalin A-Sepharose |
| title_fullStr |
Trypanosoma cruzi: Partial characterization of minor cruzipain isoforms non-adsorbed to Concanavalin A-Sepharose |
| title_full_unstemmed |
Trypanosoma cruzi: Partial characterization of minor cruzipain isoforms non-adsorbed to Concanavalin A-Sepharose |
| title_sort |
trypanosoma cruzi: partial characterization of minor cruzipain isoforms non-adsorbed to concanavalin a-sepharose |
| url |
http://hdl.handle.net/20.500.12110/paper_00144894_v104_n3-4_p122_Duschak |
| work_keys_str_mv |
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