Isolation of the Photoactive Reaction Center Complex that Contains Three Types of Fe-S Centers and a Cytochrome <italic>c</italic> Subunit from the Green Sulfur Bacterium <italic>Chlorobium limicola</italic> f. <italic>thiosulfatophilum</italic>, Strain Larsen

Abstract

The photoactive reaction center (RC) complex from the green sulfur bacterium Chlorobium limicola f. thiosulfatophilum, strain Larsen, was isolated after solubilization and ammonium sulfate fractionation followed by ion-exchange chromatography. The spectrum of the complex was almost identical with that of the similar RC complex isolated by Feiler et al. [(1992) Biochemistry 31: 2608-2614] except for the presence of cytochrome c5il instead of c553 in the latter study. A molecular ratio of BChl a to P840 of the isolated RC complex was assayed to be 25-35. SDSPAGE analysis revealed that the isolated complex contained three major polypeptides with apparent molecular masses of 68, 41 and 21 kDa, respectively. The 21-kDa polypeptide was identified to be a heme-binding protein by staining the gel for peroxidase activity. The cytochrome cssl was oxidized by flash light in a biphasic manner with half times of 90 and 390 /is, respectively, that coincided with the reduction half times of P840+. Three distinct iron-sulfur centers assigned to FA, FB and Fx, respectively, from their g-values were detected by EPR spectroscopy at cryogenic temperature. These results suggest that the present preparation contains a minimal functional unit of the RC of this bacterium, and that this complex appears to lie on a evolutionary line between RC's of purple bacteria and photosystem I.