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Memórias do Instituto Oswaldo Cruz
Fundação Oswaldo Cruz, Fiocruz
ISSN: 1678-8060
EISSN: 1678-8060
Vol. 93, No. 2, 1998, pp. 219-224
Bioline Code: oc98042
Full paper language: English
Document type: Research Article
Document available free of charge

Memórias do Instituto Oswaldo Cruz, Vol. 93, No. 2, 1998, pp. 219-224

 en Purification and partial characterization of Trypanosoma cruzi triosephosphate isomerase
SC Bourguignon; MN Meirelles; RS Pacheco & S Giovanni De Simone

Abstract

The enzyme triosephosphate isomerase (TPI, EC 5.3.1.1) was purified from
extracts of epimastigote forms of Trypanosoma cruzi. The
purification steps included: hydrophobic interaction chromatography on
phenyl-Sepharose, CM-Sepharose, and high performance liquid gel filtration
chromatography. The CM-Sepharose material contained two bands (27 and 25
kDa) with similar isoelectric points (pI 9.3-9.5) which could be separated
by gel filtration in high performance liquid chromatography. Polyclonal
antibodies raised against the porcine TPI detected one single polypeptide
on western blot with a molecular weight (27 kDa) identical to that purified
from T. cruzi. These antibodies also recognized only one band of
identical molecular weight in western blots of several other
trypanosomatids (Blastocrithidia culicis, Crithidia desouzai,
Phytomonas serpens, Herpertomonas samuelpessoai). The presence of only
one enzymatic form of TPI in T. cruzi epimastigotes was confirmed by
agarose gel activity assay and its localization was established by
immunocytochemical analysis. The T. cruzi purified TPI (as well as
other trypanosomatid' TPIs) is a dimeric protein, composed of two identical
subunits with an approximate mw of 27,000 and it is resolved on two
dimensional gel electrophoresis with a pI of 9.3. Sequence analysis of the
N-terminal portion of the 27 kDa protein revealed a high homology to
Leishmania mexicana and T. brucei proteins.

Keywords
Trypanosoma cruzi - triosephosphate isomerase - purification - sequence

 
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