Pipecolic acid is an osmoprotectant for Escherichia coli taken up by the general osmoporters ProU and ProP

CNRS URA-256, Département Membranes et Osmorégulation, Université de Rennes I, Campus de Beaulieu, Av. du Général Leclerc, 35042 Rennes Cedex, France ABSTRACT Exogenously supplied L-pipecolic acid was accumulated by Escherichia coli cells and protected them while growing at inhibitory osmolarity. Using specific uptake mutants and competitive assays, we established that the imino acid enters the cells through the ProP and ProU systems with K m values of 225 and 53 µM, respectively. Surprisingly, in spite of the requirement for the wild-type proX gene for osmoprotective ability, no binding activity of labelled pipecolate with the periplasmic protein encoded by proX could be detected. In an attempt to demonstrate whether the two porters (ProP and ProU) are the only carriers involved in osmoregulation, a variety of molecules known for their intracellular osmolarity-dependent accumulation in various organisms were investigated. N -Dimethylproline (proline betaine), N -dimethylglycine, homobetaine (β-alanine betaine), -butyrobetaine and dimethylsulfonio-propionate were found to be capable of promoting the growth of osmotically stressed E. coli. All of these molecules enter bacterial cells via ProP and ProU porters. None of the osmoprotectants except N -dimethylproline was able to bind the periplasmic protein encoded by proX , while this protein was necessary for their uptake. Apparently, ProP and ProU are the sole osmoporters involved in osmolyte influx into E. coli cells. Author for correspondence: Carlos Blanco. Tel: +33 99286140. Fax: +33 99286700. Keywords: Escherichia coli , osmoprotection, pipecolic acid, ProP and ProU systems, periplasmic binding protein