L. Pieulle et al., ISOLATION AND CHARACTERIZATION OF THE PYRUVATE-FERREDOXIN OXIDOREDUCTASE FROM THE SULFATE-REDUCING BACTERIUM DESULFOVIBRIO-AFRICANUS, Biochimica et biophysica acta. Protein structure and molecular enzymology, 1250(1), 1995, pp. 49-59
We report the first purification and characterization of a pyruvate-fe
rredoxin oxidoreductase (FOR) from a sulfate-reducing bacterium, Desul
fovibrio africanus. The enzyme as isolated is highly stable in the pre
sence of oxygen and exhibits a specific activity of 14 U/mg. D. africa
nus POR is a 256 kDa homodimer which contains thiamine pyrophosphate (
TPP) and iron-sulfur clusters. EPR spectroscopic study of the enzyme i
ndicates the presence of three [4Fe-4S](2+/1+) centers/subunit. The mi
dpoint potentials of the three centers are -390 mV, -515 mV and -540 m
V. The catalytic mechanism of FOR involves a free radical intermediate
which disappears when coenzyme A is added. This behaviour is discusse
d in terms of an electron-transport chain from TPF to the acceptor. Th
e enzyme activated by dithioerythritol shows an exceptionally high act
ivity compared with other mesophile PORs and becomes very sensitive to
oxygen in contrast to the enzyme before activation. The comparison of
EPR spectra given by the as isolated and activated enzymes shows that
neither the nature, nor the arrangement of FeS centers are affected b
y the activation process. D. africanus ferredoxins I and II are involv
ed as the physiological electron carriers of the enzyme. FOR was shown
to be located in the cytoplasm by immunogold labelling.