Affiliation:
1. Laboratory of Molecular Biology, Department of Microbiology and Immunology, University of South Alabama College of Medicine, Mobile, Alabama 36688
Abstract
ABSTRACT
The obligate intracytoplasmic pathogen
Rickettsia prowazekii
relies on the transport of many essential compounds from the cytoplasm of the eukaryotic host cell in lieu of de novo synthesis, an evolutionary outcome undoubtedly linked to obligatory growth in this metabolite-replete niche. The paradigm for the study of rickettsial transport systems is the ATP/ADP translocase Tlc1, which exchanges bacterial ADP for host cell ATP as a source of energy, rather than as a source of adenylate. Interestingly, the
R. prowazekii
genome encodes four open reading frames that are highly homologous to the well-characterized ATP/ADP translocase Tlc1. Therefore, by annotation, the
R. prowazekii
genome encodes a total of five ATP/ADP translocases: Tlc1, Tlc2, Tlc3, Tlc4, and Tlc5. We have confirmed by quantitative reverse transcriptase PCR that mRNAs corresponding to all five
tlc
homologues are expressed in
R. prowazekii
growing in L-929 cells and have shown their heterologous protein expression in
Escherichia coli
, suggesting that none of the
tlc
genes are pseudogenes in the process of evolutionary meltdown. However, we demonstrate by heterologous expression in
E. coli
that only Tlc1 functions as an ATP/ADP transporter. A survey of nucleotides and nucleosides has determined that Tlc4 transports CTP, UTP, and GDP. Intriguingly, although GTP was not transported by Tlc4, it was an inhibitor of CTP and UTP uptake and demonstrated a
K
i
similar to that of GDP. In addition, we demonstrate that Tlc5 transports GTP and GDP. We postulate that Tlc4 and Tlc5 serve the primary function of maintaining intracellular pools of nucleotides for rickettsial nucleic acid biosynthesis and do not provide the cell with nucleoside triphosphates as an energy source, as is the case for Tlc1. Although heterologous expression of Tlc2 and Tlc3 was observed in
E. coli
, we were unable to identify substrates for these proteins.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Cited by
58 articles.
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