Complementation of a reaction center-deficient Rhodobacter sphaeroides pufLMX deletion strain in trans with pufBALM does not restore the photosynthesis-positive phenotype

Abstract

The puf operon in Rhodobacter sphaeroides is composed of the genes for the photosynthetic reaction center L and M subunits, light-harvesting antenna complex I, and one other open reading frame termed pufX. Complementation of a reaction center-deficient, photosynthetically incompetent pufLMX deletion strain in trans with a fragment containing the entire puf operon, including pufX and an additional 1,100 base pairs of DNA downstream of pufX, restored the reaction center and the photosynthesis-positive phenotype. Complementation of the same strain with pufBALM restores the reaction center to the level seen with the entire puf operon but not the photosynthesis-positive phenotype. Northern (RNA) blot analysis revealed that oxygen regulated transcription was not blocked in the absence of pufX and the downstream region. Spectroscopic and protein analyses indicated that the pigment-binding protein complexes, including the reaction center, were expressed and showed normal absorption characteristics. A 20% reduction in the amount of light-harvesting antenna complex II and a corresponding increase in the amount of light-harvesting antenna complex I were observed in the deletion strain harboring the plasmid with the puf insert lacking the pufX gene and the downstream region compared with those complemented with the entire puf operon and an additional downstream 1,100 base pairs.