Synthesis of pyruvate: ferredoxin oxidoreductase and alcohol dehydrogenase E enzymes during Giardia intestinalis excystation.
BACKGROUND
Giardia intestinalis is a unicellular parasite of worldwide distribution. It causes an intestinal illness known as giardiasis, and it is probably the earliest diverging eukaryotic microorganism. Previously, changes have been reported in the expression of mRNAs at several stages of the life cycle; however specific enzymatic activity changes have not been explored.


OBJECTIVE
The expression of pyruvate ferredoxin oxidoreductase (PFOR) and alcohol dehydrogenase E (ADHE) enzymes was measured in cyst and trophozoite stages, and during the excystation process.


METHODS
Recombinant proteins were generated for PFOR and ADHE to be used as antigens in the production of polyclonal antibodies for the detection of native proteins by Western Blot. The enzymatic activity of ADHE and glutamate dehydrogenase (GDH) was evaluated by spectrophotometric assays.


RESULTS
PFOR (139 kDa) and ADHE (97 kDa) proteins were detected in trophozoites, but not in cysts. During excystation, ADHE protein was detected after the first phase of induction, but the PFOR protein appeared only after the second phase. This indicated that both proteins were synthesized during excystation, although at different times. ADHE enzymatic activity was present only in trophozoites and not in cysts whereas GDH activity was detected in both stages.


CONCLUSIONS
These results conclusively showed that PFOR and ADHE enzymes were translated during the excystation process and is strong evidence that active protein synthesis was occurring during excystation.
Niño
Carlos Alberto
CA
Laboratorio de investigaciones Básicas en Bioquímica, Departamento de Química, Universidad Nacional de Colombia, Bogotá, DC, Colombia.
Wasserman
Moisés
M
D000426:Alcohol Dehydrogenase / Q000096:biosynthesis*; D016829:Giardia lamblia / Q000201:enzymology* / Q000254:growth & development*; D050743:Pyruvate Synthase / Q000096:biosynthesis*
D000426:Alcohol Dehydrogenase; D050743:Pyruvate Synthase