Molecular and biochemical characterization of hexokinase from Trypanosoma cruzi.
Mol Biochem Parasitol
; 126(2): 251-62, 2003 Feb.
Article
em En
| MEDLINE
| ID: mdl-12615324
The Trypanosoma cruzi hexokinase gene has been cloned, sequenced, and expressed as an active enzyme in Escherichia coli. Sequence analysis revealed 67% identity with its counterpart in Trypanosoma brucei but low similarity with all other available hexokinase sequences including those of human. It contains an N-terminal peroxisome-targeting signal (PTS-2) and has a calculated basic isoelectric point (pI = 9.67), a feature often associated with glycosomal proteins. The polypeptide has a predicted mass of approximately 50 kDa similar to that of many non-vertebrate hexokinases and the vertebrate hexokinase isoenzyme IV. The natural enzyme was purified to homogeneity from T. cruzi epimastigotes and appeared to exist in several aggregation states, an apparent tetramer being the predominant form. Its kinetic properties were compared with those of the purified recombinant protein. Higher K(m) values for glucose and ATP were found for the (His)(6)-tag-containing recombinant hexokinase. However, removal of the tag produced an enzyme displaying similar values as the natural enzyme (K(m) for glucose = 43 and 60 microM for the natural and the recombinant protein, respectively). None of these enzymes presented activity with fructose. As reported previously for hexokinases from several trypanosomatids, no inhibition was exerted by glucose 6-phosphate (G6-P). In contrast, a mixed-type inhibition was observed with inorganic pyrophosphate (PPi, K(i) = 0.5mM).
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Trypanosoma cruzi
/
Hexoquinase
Limite:
Animals
/
Humans
Idioma:
En
Revista:
Mol Biochem Parasitol
Ano de publicação:
2003
Tipo de documento:
Article
País de afiliação:
Venezuela
País de publicação:
Holanda