A PURIFIED FERREDOXIN FROM GIARDIA-DUODENALIS

A ferredoxin has been purified to homogeneity from the ancient protozo an parasite Giardia duodenalis. As far as we know, this is the first e lectron transport protein to be characterised from the organism. The f erredoxin exhibits absorption maxima at 296 and 406 nm with molar abso rption coefficients of epsilon(296) = 16650 +/- 240 M(-1) cm(-1) and e psilon(406) = 13100 +/- 370 M(-1) cm(-1) respectively. The A(406)/A(29 6) ratio ranged over 0.78-0.82. The molecular mass of the apoprotein c alculated by mass spectrometry was 5730 +/- 100 Da and the minimum mol ecular mass by amino acid analysis was 5926Da. There were four cystein e residues/molecule protein but no methionine, arginine, histidine or tyrosine. The absence of these latter residues is consistent with the amino acid content of most ferredoxins. The N-terminal amino acid sequ ence exhibited greatest similarity to Desulfovibrio gigas ferredoxin I I and indicated the potential to coordinate an iron-sulfur cluster. Th ere were 3.21 +/- 0.41 mol sulfide and 2.65 +/- 0.06 mol iron/mol prot ein. Electron paramagnetic resonance studies of this protein have indi cated the presence of an iron-sulfur centre consistent with those of k nown ferredoxins. Ferredoxin serves as a biological electron acceptor from giardial pyruvate dehydrogenase with metronidazole as a terminal electron acceptor. Such a pathway may serve as a possible mechanism fo r the reductive activation of metronidazole in this parasite. A second ferredoxin has been purified to homogeneity, but at this stage there is insufficient material to fully characterise this protein. No other low-molecular-mass electron transport proteins have been identified in Giardia under the growth conditions described.