Molecular insights into FucR transcription factor to control the metabolism of -fucose in Bifidobacterium longum subsp. infantis.

Bifidobacterium longum subsp. infantis commonly colonizes the human gut and is capable of metabolizing fucose, which is abundant in the gut. Multiple studies have focused on the mechanisms of -fucose utilization by B. longum subsp. infantis, but the regulatory pathways governing the expression of these catabolic processes are still unclear. In this study, we have conducted a structural and functional analysis of fucose metabolism transcription factor FucR derived from B. longum subsp. infantis Bi-26. Our results indicated that FucR is a fucose-sensitive repressor with more α-helices, fewer β-sheets, and β-turns. Transcriptional analysis revealed that FucR displays weak negative self-regulation, which is counteracted in the presence of -fucose. Isothermal titration calorimetry indicated that FucR has a 2:1 stoichiometry with fucose. The key amino acid residues for FucR binding fucose are Asp280 and Arg331, with mutation of Asp280 to Ala resulting in a decrease in the affinity between FucR and fucose with the K value from 2.58 to 11.68 μM, and mutation of Arg331 to Ala abolishes the binding ability of FucR towards fucose. FucR specifically recognized and bound to a 20-bp incomplete palindrome sequence (5'-ACCCCAATTACGAAAATTTTT-3'), and the affinity of the fucose-loaded FucR for the DNA fragment was lower than apo-FucR. The results provided new insights into the regulating -fucose metabolism by B. longum subsp. infantis.