DsdX Is the Second d -Serine Transporter in Uropathogenic Escherichia coli Clinical Isolate CFT073

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Abstract
D -Serine is an amino acid present in mammalian urine that is inhibitory to Escherichia coli strains lacking a functional dsdA gene. Counterintuitively, a dsdA strain of E. coli clinical isolate CFT073 hypercolonizes the bladder and kidneys of mice relative to wild type during a coinfection in the murine model of urinary tract infection. We are interested in the mechanisms for uptake of d -serine in CFT073. d -Serine enters E. coli K-12 via CycA, the d -alanine transporter and d -cycloserine sensitivity locus. CFT073 cycA can grow on minimal medium with d -serine as a sole carbon source. The dsdX gene of the dsdCXA locus is a likely candidate for an additional d -serine transporter based on its predicted amino acid sequence similarity to gluconate transporters. In minimal medium, CFT073 dsdX can grow on d -serine as a sole carbon source; however, CFT073 dsdX cycA cannot. Additionally, CFT073 dsdXA cycA is not sensitive to inhibitory concentrations of d -serine during growth on glycerol and d -serine minimal medium. d -[ 14 C]serine uptake experiments with CFT073 dsdX cycA harboring dsdX or cycA recombinant plasmids confirm that d -serine is able to enter E. coli cells via CycA or DsdX. In whole-cell d -[ 14 C]serine uptake experiments, DsdX has an apparent K m of 58.75 μM and a V max of 75.96 nmol/min/mg, and CycA has an apparent K m of 82.40 μM and a V max of 58.90 nmol/min/mg. Only d -threonine marginally inhibits DsdX-mediated d -serine transport, whereas d -alanine, glycine, and d -cycloserine inhibit CycA-mediated d -serine transport. DsdX or CycA is sufficient to transport physiological quantities of d -serine, but DsdX is a d -serine-specific permease.