Microorganisms exhibit unparalleled capabilities for the consumption of naturally occurring and man-made sources of carbon. Their impressive ability to consume inert aromatic compounds is critical for environmental carbon cycling and has major implications for bioremediation, alternative energy and sustainable production of chemical feedstocks (1
). Much of the microbial catabolism of aromatic compounds is related to lignin, a highly abundant polymer that is one of the components of plant biomass (2
). A central pathway for the consumption of the lignin-derived aromatic compounds is the β-ketoadipate pathway. In this pathway, protocatechuate (3,4-dihydroxybenzoate; referred to hereafter as PCA) and catechol are converted into the eponymous β-ketoadipate and, ultimately, acetyl-coenzyme and succinyl-coenzyme A (4
). The fact that these products can be converted anabolically into triglyceride precursors of biodiesel or into high-value compounds like polyketide antibiotics has motivated much renewed interest in this pathway (5
In addition to being a prototype for the catabolism of lignin-derived aromatic compounds, the β-ketoadipate pathway has been a model system for studies of how microorganisms regulate the catabolism of aromatic compounds at the genetic level (4
). The theme that has emerged from the investigations by multiple groups is that genes encoding enzymes of the pathway are regulated by either LysR or IclR family transcription factors (4
). Mostly, these transcription factors mediate environmental surveillance as receptors for aromatic ligands that modulate their DNA-binding ability. Our recent studies of aromatic catabolism in Streptomyces
bacteria resulted in the discovery of a MarR family transcription factor called PcaV that regulates genes encoding enzymes of the PCA branch of the β-ketoadipate pathway (14
). Beyond its regulation of a central pathway for aromatic catabolism, PcaV is of interest because it is the only known member of the MarR family that regulates the β-ketoadipate pathway.
The MarR family of transcription factors is a large group of proteins encoded by >12 000 genes in the publicly available genomes of bacteria and archaea. While these proteins can be either transcriptional repressors or activators, they have been ascribed roles in controlling the expression of genes underlying catabolic pathways, stress responses, virulence and multi-drug resistance (17–21
). To date, the physiological roles of ~100 of these proteins have been characterized in detail (22
). While a subset of MarR family members regulate adaptive responses to oxidative stress through the formation of disulfide bonds that influence DNA binding (23–27
), the majority of these proteins regulate gene expression through ligand-mediated attenuation of DNA binding. Our understanding of the molecular mechanism of regulation by ligand-responsive MarR family proteins is limited because the identity of the ligand is often unknown (22
). Further, in most cases wherein structures of MarR family members in complex with ligands have been reported, the ligand’s physiological role cannot be easily connected to the functions of the regulated genes (22
). As MarR family members play important roles in antibiotic resistance, virulence and catabolism, studies of their molecular mechanisms have implications for medicine and biotechnology.
Our discovery that PCA regulates the PcaV-dependent transcriptional activation of the corresponding structural genes in Streptomyces coelicolor provided a unique opportunity to study how a MarR family transcription factor responds to its natural ligand. Bioinformatics, electrophoretic mobility shift assays (EMSAs), mutagenesis, isothermal calorimetry (ITC) and in vivo transcription assays were used to elucidate the regulatory mechanism of PcaV. Further, we report the crystal structures of apo-PcaV and PcaV bound to its ligand PCA. Our findings are particularly noteworthy because they provide a contrast to the few crystal structures of MarR family members bound to ligands whose identities cannot be connected to the functions of the genes that they regulate and, as a consequence, new insights into the molecular basis of the transcriptional regulation of MarR proteins by their native ligands.