Plasticity in oligomerization, operator architecture, and DNA binding in the mode of action of a bacterial B12-based photoreceptor

Newly discovered bacterial photoreceptors called CarH sense light by using 5-deoxyadenosylcobalamin (AdoCbl). They repress their own expression and that of genes for carotenoid synthesis by binding in the dark to operator DNA as AdoCbl-bound tetramers, whose light-induced disassembly relieves repression. High-resolution structures of Thermus thermophilus CarH(Tt) have provided snapshots of the dark and light states and have revealed a unique DNA-binding mode whereby only three of four DNA-binding domains contact an operator comprising three tandem direct repeats. To gain further insights into CarH photoreceptors and employing biochemical, spectroscopic, mutational, and computational analyses, here we investigated CarH(Bm) from Bacillus megaterium. We found that apoCarH(Bm), unlike monomeric apoCarH(Tt), is an oligomeric molten globule that forms DNA-binding tetramers in the dark only upon AdoCbl binding, which requires a conserved W-X-9-EH motif. Light relieved DNA binding by disrupting CarH(Bm) tetramers to dimers, rather than to monomers as with CarH(Tt). CarH(Bm) operators resembled that of CarH(Tt), but were larger by one repeat and overlapped with the -35 or -10 promoter elements. This design persisted in a six-repeat, multipartite operator we discovered upstream of a gene encoding an Spx global redox-response regulator whose photoregulated expression links photooxidative and general redox responses in B. megaterium. Interestingly, CarH(Bm) recognized the smaller CarH(Tt) operator, revealing an adaptability possibly related to the linker bridging the DNA- and AdoCbl-binding domains. Our findings highlight a remarkable plasticity in the mode of action of B-12-based CarH photoreceptors, important for their biological functions and development as optogenetic tools.