Paracoccus denitrificans possesses two BioR homologs having a role in regulation of biotin metabolism

Recently, we determined that BioR, the GntR family of transcription factor, acts
as a repressor for biotin metabolism exclusively distributed in certain species of
a-proteobacteria, including the zoonotic agent Brucella melitensis and the plant
pathogen Agrobacterium tumefaciens. However, the scenario is unusual in Paracoccus
denitrificans, another closely related member of the same phylum a-proteobacteria
featuring with denitrification. Not only does it encode two BioR
homologs Pden_1431 and Pden_2922 (designated as BioR1 and BioR2, respectively),
but also has six predictive BioR-recognizable sites (the two bioR homolog
each has one site, whereas the two bio operons (bioBFDAGC and bioYB) each contains
two tandem BioR boxes). It raised the possibility that unexpected complexity
is present in BioR-mediated biotin regulation. Here we report that this is the
case. The identity of the purified BioR proteins (BioR1 and BioR2) was confirmed
with LC-QToF-MS. Phylogenetic analyses combined with GC percentage raised a
possibility that the bioR2 gene might be acquired by horizontal gene transfer. Gel
shift assays revealed that the predicted BioR-binding sites are functional for the
two BioR homologs, in much similarity to the scenario seen with the BioR site of
A. tumefaciens bioBFDAZ. Using the A. tumefaciens reporter system carrying a
plasmid-borne LacZ fusion, we revealed that the two homologs of P. denitrificans
BioR are functional repressors for biotin metabolism. As anticipated, not only
does the addition of exogenous biotin stimulate efficiently the expression of bioYB
operon encoding biotin transport/uptake system BioY, but also inhibits the transcription
of the bioBFDAGC operon resembling the de novo biotin synthetic pathway.
EMSA-based screening failed to demonstrate that the biotin-related
metabolite is involved in BioR-DNA interplay, which is consistent with our former
observation with Brucella BioR. Our finding defined a complex regulatory
network for biotin metabolism in P. denitrificans by two BioR proteins.