The pair of fatty acid degradation regulator (FadR)-characterized pseudo-palindromic motifs spacing with 11 bp contribute unequally to the DNA binding ability of the HTH-type transcriptional regulator Rv0494 from Mycobacterium tuberculosis H37Rv.

Abstract

Transcriptional regulation is an effective way for bacteria to respond to various environmental challenges, such as carbon source scarcity. Acting as a transcription factor belonging to the fatty acid degradation regulator (FadR) subfamily, the HTH-type transcriptional regulator Rv0494 is reported to function in persistence, starvation response and regulation of very-long-chain fatty acid synthesis in Mycobacterium tuberculosis (Mtu), a notorious pathogen that causes tuberculosis. However, DNA sequence dependence of Rv0494, especially the contributions of the pair of FadR-characterized palindromic motifs spaced with 11 bp in the upstream sequence of rv0494 to the DNA binding ability of Rv0494, remains unclear. Through structural biology and biochemical approaches, this study demonstrated that although Rv0494 binds to DNA in a FadR-conserved way, such as the conserved key residues responsible for DNA binding and the prevalent functional homodimer, the pair of FadR-characterized pseudo-palindromic motifs spaced by 11 bp contribute unequally to the DNA binding activity of Rv0494, probably due to an additional DNA binding activity contributed by a previously unidentified palindromic sequence with a spacer of 5 bp, which is composed of the major-contributing pseudo-palindromic motif and a complementary sequence separated from it by 5 bp. This asymmetric motif dependence is further validated in the upstream sequences of more carbon metabolism-related genes, such as rv0467 (icl1) that encodes isocitrate lyase. However, binding does not necessarily lead to transcriptional repression. This work systematically characterizes the sequence dependence behavior of Rv0494, laying the foundation for further elucidating the underlying molecular mechanisms.