Aerobic and Anaerobic Mg-Protoporphyrin Monomethyl Ester Cyclases in Purple Bacteria: A STRATEGY ADOPTED TO BYPASS THE REPRESSIVE OXYGEN CONTROL SYSTEM

Abstract

Two different mechanisms for Mg-protoporphyrin monomethyl ester (MgPMe) cyclization are shown to coexist in Rubrivivax gelatinosus and are proposed to be conserved in all facultative aerobic phototrophs: an anaerobic mechanism active under photosynthesis or low oxygenation, and an aerobic mechanism active only under high oxygenation conditions. This was confirmed by analyzing the bacteriochlorophyll accumulation in the wild type and in three mutant strains grown under low or high aeration. A mutant lacking the acsF gene is photosynthetic, exhibits normal bacteriochlorophyll accumulation under low oxygenation and anaerobiosis, and accumulates MgPMe under high oxygenation. The photosynthesis-deficient bchE mutant produces bacteriochlorophyll only under high oxygenation and accumulates MgPMe under low oxygenation and anaerobiosis. The double knockout mutant is devoid of photosystem and accumulates MgPMe under both conditions indicating the involvement of the two enzymes at the same step of the biosynthesis pathway. Oxygen-mediated expression of bchE was studied in the wild type and in a regulatory mutant. The reverse transcriptase-PCR and the bchE promoter activity results demonstrate that the expression of the bchE gene is oxygen-independent and suggest that it is rather the enzyme activity that should be oxygen-sensitive. No obvious sequence similarities were found between oxygen-dependent AcsF and the oxygen-independent anaerobic Mg-protoporphyrin monomethylester cyclase (BchE) enzymes. However, common to all BchE proteins is the conserved CXXX-CXXC sequence. This motif is essential for 4Fe-4S cluster formation in many anaerobic enzymes. Expression and purification of BchE were achieved, and the UV-visible spectral analyses confirmed the presence of an active 4Fe-4S cluster in this protein. The use of different classes of enzymes catalyzing the same reaction under different oxygen growth conditions appears to be a common feature of different biosynthetic pathways, and the benefit of possessing both aerobic and anaerobic systems is discussed.