Chlamydia trachomatis Transports NAD via the Npt1 ATP/ADP Translocase

Abstract

Obligate intracellular bacteria comprising the order Chlamydiales lack the ability to synthesize nucleotides de novo and must acquire these essential compounds from the cytosol of the host cell. The environmental protozoan endosymbiont Protochlamydia amoebophila UWE25 encodes five nucleotide transporters with specificities for different nucleotide substrates, including ATP, GTP, CTP, UTP, and NAD. In contrast, the human pathogen Chlamydia trachomatis encodes only two nucleotide transporters, the ATP/ADP translocase C. trachomatis Npt1 (Npt1 Ct ) and the nucleotide uniporter Npt2 Ct , which transports GTP, UTP, CTP, and ATP. The notable absence of a NAD transporter, coupled with the lack of alternative nucleotide transporters on the basis of bioinformatic analysis of multiple C. trachomatis genomes, led us to re-evaluate the previously characterized transport properties of Npt1 Ct . Using [adenylate-32 P]NAD, we demonstrate that Npt1 Ct expressed in Escherichia coli enables the transport of NAD with an apparent K m and V max of 1.7 M and 5.8 nM mg ؊1 h ؊1 , respectively. The K m for NAD transport is comparable to the K m for ATP transport of 2.2 M, as evaluated in this study. Efflux and substrate competition assays demonstrate that NAD is a preferred substrate of Npt1 Ct compared to ATP. These results suggest that during reductive evolution, the pathogenic chlamydiae lost individual nucleotide transporters, in contrast to their environmental endosymbiont relatives, without compromising their ability to obtain nucleotides from the host cytosol through relaxation of transport specificity. The novel properties of Npt1 Ct and its conservation in chlamydiae make it a potential target for the development of antimicrobial compounds and a model for studying the evolution of transport specificity.

Benzylidene Acylhydrazides Inhibit Chlamydial Growth in a Type III Secretion- and Iron Chelation-Independent Manner

“…ATP uptake by E. coli ATM1173, which expresses His-tagged ATP/ADP translocase (CTL0321), was determined as previously reported (36) with modifications. In preliminary experiments, ATM1172, which carries the empty pET19 plasmid, failed to practically take up radioactivity (data not shown), which is consistent with the findings of Fisher et al (36). Therefore, ATM1172 was not included in later experiments.…”

“…As expected, NAD, which is transported into chlamydiae through the same translocase (36), inhibited the uptake of [␣-…”

Chlamydia cell biology and pathogenesis

“…Chlamydiae scavenge nutrients from various sources, for example, from the lysosomal-mediated degradation of proteins 7 , using various transporters 53,70 . Although chlamydiae can synthesize common bacterial lipids, the membrane of C. trachomatis contains eukaryotic lipids, including phosphatidylcholine, phosphatidylinositol, sphingomyelin and cholesterol 21 .…”

CTL0511 from Chlamydia trachomatis Is a Type 2C Protein Phosphatase with Broad Substrate Specificity

“…Broadening of the substrate specificity of the chlamydial Hanks'-type kinases and Ser/Thr protein phosphatase to allow for interactions with (P-)Tyr may have allowed for the loss of PTPs and BY kinases during reductive evolution of the chlamydial genomes. Such a pathway would mirror the evolution of the chlamydial nucleotide transporters, which are fewer and have broader substrate preferences than the chlamydial ancestors (68,69). In light of the broad substrate specificities of CTL0511 and the C. pneumoniae PknD, the tyrosine phosphorylation capabilities of PknD and Pkn1 from C. trachomatis should be reassessed.…”