Plasticity in Repressor-DNA Interactions Neutralizes Loss of Symmetry in Bipartite Operators

Abstract

Transcription factor-DNA interactions are central to gene regulation. Many transcription factors regulate multiple target genes and can bind sequences that do not conform strictly to the consensus. To understand the structural mechanism utilized by the transcription regulators to bind diverse target sequences, we have employed the repressor AraR from Bacillus subtilis as a model system. AraR is known to bind to eight different operator sites in the bacterial genome. Although there are differences in the sequences of four of these operators, ORE1, ORX1, ORA1, and ORR3, the AraR-DNA binding domain (AraR-DBD) as well as full-length AraR unexpectedly binds to each of these sequences with similar affinities as measured by fluorescence anisotropy experiments. We have determined crystal structures of AraR-DBD in complex with two different natural operators ORE1 and ORX1 up to 2.07 and 1.97 Å resolution, respectively. These structures were compared with the previously reported structures of AraR-DBD bound to two other natural operators (ORA1 and ORR3). Interactions of two molecules of AraR-DBD with the symmetric operator, ORE1, are identical, but their interaction with the non-symmetric operator ORX1 results in breakdown of the symmetry in protein-DNA interactions. The novel interactions observed are accompanied by local conformational change in the DNA. ChIP-sequencing (ChIP-Seq) data on other transcription factors has shown that they can bind to diverse targets, and hence the plasticity exhibited by AraR may be a general phenomenon. The ability of transcription factors to form alternate interactions may be important for employment in new functions and evolution of novel regulatory circuits.The binding of proteins to cognate DNA sequences with high specificity is critical for a number of cellular processes. Among such proteins, the ability to detect and bind target sequences present at different locations in the whole genome is especially crucial for the function of transcription factors. This ability enables the transcriptional regulatory pathways to function with remarkable precision and sensitivity. The specificity of transcription factor for its cognate DNA sequence is largely achieved due to non-covalent interactions between the amino acid side chains and the functional groups present on the DNA bases in the major and minor groove (1). In addition, the shape of the DNA also plays a significant role in specific protein-DNA recognition (2-5). The availability of transcription factor binding data spanning entire genomes has led to identification of new targets for transcription factors. It has been observed that a single transcription factor can bind to several different sites throughout the genome. Sequence deviations have been seen in target sites that are recognized by the same transcription factor. Examples include glp repressors of Escherichia coli and NdgR from Streptomyces coelicolor, which recognize 13 and 19 different operator sequences, respectively (6, 7). The sequence differences suggest the possibility o...

Radiation-Induced Effect on Spin-Selective Electron Transfer through Self-Assembled Monolayers of ds-DNA

“…To study the effect of ionizing radiation on ds-DNA, we took a 21-base DNA sequence of the operator DNA (ORR3) that binds specifically to the protein AraR from Bacillus subtilis [21]. SAM of ds-DNA was prepared on Ni/Au thin film and subsequently exposed to γ-radiation at different time scales.…”