Molecular mechanisms of xylose utilization by Pseudomonas fluorescens: overlapping genetic responses to xylose, xylulose, ribose and mannitol.

Bacterial degradation of xylose is sequentially mediated by two enzymes - an isomerase (XutA) and a xylulokinase (XutB) - with xylulose as an intermediate. Pseudomonas fluorescens SBW25, though capable of growth on xylose as a sole carbon source, encodes only one degradative enzyme XutA at the xylose utilization (xut) locus. Here, using site-directed mutagenesis and transcriptional assays, we have identified two functional xylulokinase-encoding genes (xutB1 and xutB2) and further show that expression of xutB1 is specifically induced by xylose. Surprisingly, xylose-induced xutB1 expression is mediated by the mannitol-responsive regulator MtlR, using xylulose rather than xylose as the direct inducer. In contrast, expression of the xutA operon is regulated by XutR - a transcriptional activator of the AraC family - in a xylose-, xylulose- and ribose-dependent manner. Detailed genetic and biochemical analyses of XutR, including DNase I footprinting assays, suggest an unconventional model of XutR regulation that does not involve DNA-looping, a mechanism typically found for AraC-type regulators from enteric bacteria. XutR functions as a dimer and recognizes two inverted repeat sequences, but binding to one half site is weak thus requiring an inducer molecule such as xylose for activation.
(© 2015 John Wiley & Sons Ltd.)