Functional Identification of the Hypoxanthine/Guanine Transporters YjcD and YgfQ and the Adenine Transporters PurP and YicO of Escherichia coli K-12

Παπακώστας, Κωνσταντίνος; Botou, Maria; Frillingos, Stathis

doi:10.1074/jbc.m113.523340

Cited by 51 publications

(133 citation statements)

References 39 publications

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“…As a result, homology modeling has been used as a reasonable alternative in cases where no x-ray structure is available (23,48,59,61,(67)(68)(69)(70)(71)(72)(73). Using this approach, combined with induced fit docking and a detailed characterization of the specificity profile, we identified 9 residues that are important for PrnB function, Gly 56 Table 5).…”

Section: Discussionmentioning

confidence: 99%

The Aspergillus nidulans Proline Permease as a Model for Understanding the Factors Determining Substrate Binding and Specificity of Fungal Amino Acid Transporters

Gournas

Evangelidis

Αθανασόπουλος

et al. 2015

Journal of Biological Chemistry

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Background: PrnB, different from Put4p, its Saccharomyces cerevisiae orthologue, is a highly specific L-proline transporter of Aspergillus nidulans. Results: Identification of 12 residues important for PrnB activity and/or specificity. Put4p-mimicking mutants of PrnB recognize other Put4p substrates. Conclusion: Residues variable in the yeast amino acid transporter (YAT) family determine transporter specificity. Significance: Understanding the molecular mechanisms underlying amino acid recognition and translocation through YATs.

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Section: Discussionmentioning

confidence: 99%

The Aspergillus nidulans Proline Permease as a Model for Understanding the Factors Determining Substrate Binding and Specificity of Fungal Amino Acid Transporters

Gournas

Evangelidis

Αθανασόπουλος

et al. 2015

Journal of Biological Chemistry

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“…Briefly, to initiate the metabolism of the starved spirochetes, the bacteria were preincubated at 37°C for 30 min in 1-ml reaction volumes containing HN buffer (50 mM HEPES and 50 mM NaCl, pH 7.4), 6 mM glucose, and 1.4% rabbit serum (Pelfreez). This amount of rabbit serum was estimated to contain ϳ90 nM hypoxanthine and ϳ5 nM adenine (19,20) 15 Ci/mmol) (PerkinElmer), and samples were analyzed at 10, 20, 30, 45, and 60 s following the addition of radioactivity. The rates of purine uptake (fmol/5 ϫ 10 7 spirochetes/s) were estimated for each purine concentration by linear regression, and the K m values were determined using GraphPad Prism, version 5.0.…”

Section: Methodsmentioning

confidence: 99%

“…Recently, structure-function analyses of the E. coli adenine-specific permeases PurP and YicO and hypoxanthine-guanine-specific permeases YjcD and YgfQ as well as the A. nidulans adeninehypoxanthine-guanine permease AzgA identified key amino acid residues critical for purine uptake activity (13,15). Both of these studies identified an aspartic acid and a glutamic acid residue (Asp339 and Glu394 in AzgA) to be essential for the transport function for all five COG2252 family members (13,15). These residues are conserved in BBB22 and BBB23.…”

Section: Figmentioning

confidence: 99%

“…In other species, however, specific purine transport functions have been attributed to distinct proteins. For example, Escherichia coli harbors two high-affinity adenine permeases, PurP and YicO, and two highaffinity hypoxanthine-guanine permeases, YjcD and YgfQ (12,15). Similarly, Bacillus subtilis has distinct uptake systems for hypoxanthine-guanine and adenine (16,17).…”

mentioning

confidence: 99%

“…The bbb22 and bbb23 genes encode proteins within cluster COG2252 of the nucleobase cation symporter-2 superfamily (NCS2) (7), which includes permeases found in bacteria, fungi, and plants that are specific for adenine, hypoxanthine, and/or guanine (10)(11)(12)(13)(14)(15)(16). The Aspergillus nidulans AzgA transporter, the founding member of this family of transporters, has specificity for adenine, hypoxanthine, and guanine (10,11,13).…”

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confidence: 99%

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Molecular Dissection of a Borrelia burgdorferi In Vivo Essential Purine Transport System

2015

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The Lyme disease spirochete Borrelia burgdorferi is dependent on purine salvage from the host environment for survival. The genes bbb22 and bbb23 encode purine permeases that are essential for B. burgdorferi mouse infectivity. We now demonstrate the unique contributions of each of these genes to purine transport and murine infection. The affinities of spirochetes carrying bbb22 alone for hypoxanthine and adenine were similar to those of spirochetes carrying both genes. Spirochetes carrying bbb22 alone were able to achieve wild-type levels of adenine saturation but not hypoxanthine saturation, suggesting that maximal hypoxanthine uptake requires the presence of bbb23. Moreover, the purine transport activity conferred by bbb22 was dependent on an additional distal transcriptional start site located within the bbb23 open reading frame. The initial rates of uptake of hypoxanthine and adenine by spirochetes carrying bbb23 alone were below the level of detection. However, these spirochetes demonstrated a measurable increase in hypoxanthine uptake over a 30-min time course. Our findings indicate that bbb22-dependent adenine transport is essential for B. burgdorferi survival in mice. The bbb23 gene was dispensable for B. burgdorferi mouse infectivity, yet its presence was required along with that of bbb22 for B. burgdorferi to achieve maximal spirochete loads in infected mouse tissues. These data demonstrate that both genes, bbb22 and bbb23, are critical for B. burgdorferi to achieve wild-type infection of mice and that the differences in the capabilities of the two transporters may reflect distinct purine salvage needs that the spirochete encounters throughout its natural infectious cycle. P urine nucleobases are required for the synthesis of DNA and RNA. Consequently, purine biosynthesis and/or transport is a critical process across all kingdoms of life. Moreover, nucleobase transporters represent possible therapeutic targets for cancer and infectious diseases (1).Borrelia burgdorferi, the causative agent of Lyme disease, has a limited genome and lacks the enzymes required for de novo purine biosynthesis (2-6). Therefore, purine salvage from host environments is important for B. burgdorferi survival and pathogenesis (7-9). Recently, our laboratory established that the genes bbb22 and bbb23, located on B. burgdorferi essential circular plasmid 26 (cp26), together encode a purine transport system that is required for hypoxanthine transport and contributes to adenine and guanine transport (7). Spirochetes lacking both bbb22 and bbb23 are noninfectious in mice (7). Conversely, these genes are dispensable for B. burgdorferi growth in nutrient-rich medium in vitro (7).The bbb22 and bbb23 genes encode proteins within cluster COG2252 of the nucleobase cation symporter-2 superfamily (NCS2) (7), which includes permeases found in bacteria, fungi, and plants that are specific for adenine, hypoxanthine, and/or guanine (10-16). The Aspergillus nidulans AzgA transporter, the founding member of this family of transporters, has specif...

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The solute transport and binding profile of a novel nucleobase cation symporter 2 from the honeybee pathogen Paenibacillus larvae

et al. 2018

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Here, we report that a novel nucleobase cation symporter 2 encoded in the genome of the honeybee bacterial pathogen Paenibacillus larvae reveals high levels of amino acid sequence similarity to the Escherichia coli and Bacillus subtilis uric acid and xanthine transporters. This transporter is named P. larvae uric acid permease‐like protein (PlUacP). Even though PlUacP displays overall amino acid sequence similarities, has common secondary structures, and shares functional motifs and functionally important amino acids with E. coli xanthine and uric acid transporters, these commonalities are insufficient to assign transport function to PlUacP. The solute transport and binding profile of PlUacP was determined by radiolabeled uptake experiments via heterologous expression in nucleobase transporter‐deficient Saccharomyces cerevisiae strains. PlUacP transports the purines adenine and guanine and the pyrimidine uracil. Hypoxanthine, xanthine, and cytosine are not transported by PlUacP, but, along with uric acid, bind in a competitive manner. PlUacP has strong affinity for adenine K m 7.04 ± 0.18 μ m , and as with other bacterial and plant NCS 2 proteins, PlUacP function is inhibited by the proton disruptor carbonyl cyanide m‐chlorophenylhydrazone. The solute transport and binding profile identifies PlUacP as a novel nucleobase transporter.

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Functional Identification of the Hypoxanthine/Guanine Transporters YjcD and YgfQ and the Adenine Transporters PurP and YicO of Escherichia coli K-12

Cited by 51 publications

References 39 publications

The Aspergillus nidulans Proline Permease as a Model for Understanding the Factors Determining Substrate Binding and Specificity of Fungal Amino Acid Transporters

The Aspergillus nidulans Proline Permease as a Model for Understanding the Factors Determining Substrate Binding and Specificity of Fungal Amino Acid Transporters

Molecular Dissection of a Borrelia burgdorferi In Vivo Essential Purine Transport System

The solute transport and binding profile of a novel nucleobase cation symporter 2 from the honeybee pathogen Paenibacillus larvae

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