Two Nucleotide Transport Proteins in Chlamydia trachomatis , One for Net Nucleoside Triphosphate Uptake and the Other for Transport of Energy

Abstract

ABSTRACT The genome of Chlamydia trachomatis , one of the most prominent human pathogens, contains two structural genes coding for proteins, herein called Npt1 Ct and Npt2 Ct (nucleoside phosphate transporters 1 and 2 of C. trachomatis ), exhibiting 68 and 61% similarity, respectively, to the ATP/ADP transporter from the intracellular bacterium Rickettsia prowazekii at the deduced amino acid level. Hydropathy analysis and sequence alignments suggested that both proteins have 12 transmembrane domains. The putative transporters were expressed as histidine-tagged proteins in Escherichia coli to study their biochemical properties. His 10 -Npt1 Ct catalyzed ATP and ADP transport in an exchange mode. The apparent K m values were 48 (ATP) and 39 (ADP) μM. ATP and ADP transport was specific since AMP, GTP, CTP, UTP, dATP, dCTP, dGTP, and dTTP did not inhibit uptake. In contrast, His 10 -Npt2 Ct transported all four ribonucleoside triphosphates with apparent K m values of 31 μM (GTP), 302 μM (UTP), 528 μM (CTP), and 1,158 μM (ATP). Ribonucleoside di- and monophosphates and deoxyribonucleotides were not substrates. The protonophore m -chlorocarbonylcyanide phenylhydrazone abolished uptake of all nucleoside triphosphates by Npt2 Ct . This observation indicated that His 10 -Npt2 Ct acts as a nucleosidetriphosphate/H + symporter energized by the proton motive force across the Escherichia coli cytoplasmic membrane. We conclude that Npt1 Ct provides chlamydiae with energy whereas Npt2 Ct catalyzes the net uptake of ribonucleoside triphosphates required for anabolic reactions.