BraG is one of the two nucleotide-binding (ATPase) subunits of LIV-I, the high-affinity, Na(+)-independent branched-chain amino acid ABC importer of Pseudomonas aeruginosa, encoded by the last gene of the braC-braD-braE-braF-braG operon. BraG is a LivF-type ABC ATPase with a Walker A ATP-binding motif. It associates peripherally with the cytoplasmic face of the inner membrane, where it pairs with BraF and binds the BraD/BraE permeases. With the periplasmic binding protein BraC, the complex couples ATP hydrolysis to the uptake of L-leucine, L-isoleucine and L-valine and, with lower affinity, L-alanine and L-threonine. All five Bra components are required for transport in reconstituted proteoliposomes.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003333 amino acid transmembrane transport | IEA GO_REF:0000117 | TOO GENERAL | Summary: True but general; the specific import processes are asserted in this review. Reason: GO:1903801 L-leucine import across plasma membrane (NEW here) and GO:1904273 L-alanine import across plasma membrane (MODIFY target here) are is_a descendants of amino acid transmembrane transport. Supporting Evidence: PMID:1400443 In this reconstituted system, leucine transport was completely dependent on the presence of all five Bra components and on ATP loaded internally to the proteoliposomes. |
| GO:0005524 ATP binding | IEA GO_REF:0000002 | ACCEPT | Summary: BraG has a canonical ABC ATPase domain with a Walker A motif. Reason: The ABC transporter domain is clear, BraF and BraG both carry ATP-binding sequences, and the reconstituted LIV-I complex consumes ATP during transport. Supporting Evidence: file:PSEAE/braG/braG-uniprot.txt Pfam; PF00005; ABC_tran; 1. PMID:2120183 both containing amino acid sequences typical of proteins with ATP-binding sites |
| GO:0005886 plasma membrane | IEA GO_REF:0000044 | ACCEPT | Summary: BraG works at the cytoplasmic face of the inner (plasma) membrane, bound to the BraD/BraE permeases. Reason: BraF and BraG were found among cell membrane proteins, and BraDEFG forms a membrane complex. As an ABC ATPase BraG is peripheral, not integral. Supporting Evidence: PMID:2120183 These proteins were found among cell membrane proteins PMID:1400443 The proteins were found to form a complex in the E. coli membrane |
| GO:0015658 branched-chain amino acid transmembrane transporter activity | IBA GO_REF:0000033 | ACCEPT | Summary: BraG contributes the ATPase to the BraCDEFG branched-chain amino acid importer. The IBD is seeded by E. coli LivF. Reason: The contributes_to qualifier is right for an ATPase subunit. LIV-I carries leucine, isoleucine and valine, and transport needs all five Bra proteins. BraG is inside the LivF clade and nothing suggests divergence. Supporting Evidence: PMID:1400443 In this reconstituted system, leucine transport was completely dependent on the presence of all five Bra components and on ATP loaded internally to the proteoliposomes. PMID:113383 The former system seemed to serve not only for the entry of leucine, isoleucine, and valine, but also for that of alanine and threonine, although less effectively. |
| GO:0015658 branched-chain amino acid transmembrane transporter activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO from the LivF/BraG ATPase signature. Same claim as the IBA row. Reason: The activity is that of the BraCDEFG complex, to which BraG is essential, so the claim holds. contributes_to would be the more accurate qualifier for a single subunit. Supporting Evidence: PMID:2120184 confirming that the braD, braE, braF, and braG genes are required for the LIV-I transport system |
| GO:0015803 branched-chain amino acid transport | IEA GO_REF:0000002 | TOO GENERAL | Summary: True but general. Reason: GO:1903801 L-leucine import across plasma membrane, asserted NEW in this review, is_a branched-chain amino acid transport. Supporting Evidence: PMID:1400443 leucine transport was completely dependent on the presence of all five Bra components |
| GO:0015804 neutral amino acid transport | IEA GO_REF:0000117 | TOO GENERAL | Summary: True but general. Reason: GO:1903801 L-leucine import across plasma membrane, asserted NEW in this review, is_a neutral amino acid transport. Supporting Evidence: PMID:1400443 leucine transport was completely dependent on the presence of all five Bra components |
| GO:0015807 L-amino acid transport | IBA GO_REF:0000033 | TOO GENERAL | Summary: L-amino acid transport IBD at the LivF node PTN000657089, seeded by BraG's own PseudoCAP L-alanine transport IMP and by E. coli LivF. Reason: The claim is true; BraG in its own WITH/FROM is expected, not circular. The node term is suitably generic for a LivF clade whose substrates are set by the partner binding proteins. For BraG itself, the specific processes GO:1903801 L-leucine import and GO:1904273 L-alanine import across plasma membrane, both asserted in this review, are is_a descendants. Supporting Evidence: PMID:1400443 Alanine and threonine in addition to branched-chain amino acids were transported by the proteoliposomes |
| GO:0015807 L-amino acid transport | IEA GO_REF:0000117 | TOO GENERAL | Summary: Same claim as the IBA row. Reason: Entailed by GO:1903801 and GO:1904273, asserted in this review. Supporting Evidence: PMID:1400443 Alanine and threonine in addition to branched-chain amino acids were transported by the proteoliposomes |
| GO:0015808 L-alanine transport | IMP PMID:18833300 High-throughput phenotypic characterization of Pseudomonas a... | MODIFY | Summary: PseudoCAP IMP seed of the PTN000657089 L-amino acid transport IBD. The PA1070 Tn5 mutant grows less well (+/-) on L-alanine as carbon source. Reason: Seed verdict SOUND. A partial growth defect is an indirect read-out, but here it agrees with direct evidence that BraCDEFG imports alanine into proteoliposomes, and BraG is required for LIV-I. braG is the last gene of the operon, so the insertion is not polar on other bra genes. The precise term is L-alanine import across plasma membrane (GO:1904273, an is_a descendant). Proposed replacements: L-alanine import across plasma membrane Supporting Evidence: PMID:18833300 In this work, mutants for each of these genes were found to be defective in Alanine metabolism PMID:1400443 Alanine and threonine in addition to branched-chain amino acids were transported by the proteoliposomes |
| GO:0016887 ATP hydrolysis activity | IEA GO_REF:0000002 | ACCEPT | Summary: BraG is an ABC ATPase subunit; the LIV-I complex hydrolyses ATP coupled to transport. Reason: This is BraG's own catalytic activity. The BraF/BraG contributions have not been separated, but both carry ABC ATPase domains. Supporting Evidence: PMID:1400443 indicating that the LIV-I transport system has an ATPase activity coupled to translocation of branched-chain amino acids across the membrane file:PSEAE/braG/braG-uniprot.txt PROSITE; PS50893; ABC_TRANSPORTER_2; 1. |
| GO:0042941 D-alanine transmembrane transport | IMP PMID:18833300 High-throughput phenotypic characterization of Pseudomonas a... | UNDECIDED | Summary: Reduced (+/-) growth of the PA1070 Tn5 mutant on D-alanine as carbon source, in a Biolog screen with plate confirmation. Reason: The phenotype is real but partial, and no uptake or binding assay was done. Nothing independent shows D-alanine to be a LIV-I substrate, so whether BraG takes part in D-alanine transport stays open (as for braF and braD). Supporting Evidence: PMID:18833300 In this work, mutants for each of these genes were found to be defective in Alanine metabolism |
| GO:1905039 carboxylic acid transmembrane transport | IEA GO_REF:0000117 | TOO GENERAL | Summary: True but general. Reason: Entailed by GO:1903801 and GO:1904273, asserted in this review. Supporting Evidence: PMID:1400443 leucine transport was completely dependent on the presence of all five Bra components |
| GO:1903801 L-leucine import across plasma membrane | IDA PMID:1400443 Solubilization and reconstitution of the Pseudomonas aerugin... | NEW | Summary: ATP-dependent leucine uptake into proteoliposomes requires BraG together with the other four Bra proteins. Reason: BraG does part of the work, supplying ATPase activity that drives translocation. Leucine is the reference LIV-I substrate. BraG has no specific substrate-import annotation at present. Supporting Evidence: PMID:1400443 In this reconstituted system, leucine transport was completely dependent on the presence of all five Bra components and on ATP loaded internally to the proteoliposomes. |
| GO:0055052 ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing | IDA PMID:1400443 Solubilization and reconstitution of the Pseudomonas aerugin... | NEW | Summary: BraG is a subunit of the BraDEFG membrane complex, which works with the periplasmic binding protein BraC. Reason: BraDEFG form a complex that can be solubilised from the membrane, and transport requires the substrate-binding protein BraC. Matches the braF review. Supporting Evidence: PMID:1400443 The proteins were found to form a complex in the E. coli membrane and solubilized from the membrane with octyl glucoside. |
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Download this section (compressed HTML)Q: Is D-alanine a genuine LIV-I substrate, or is the weak D-alanine growth defect of every bra mutant (PA1070-PA1074) indirect?
Experiment: Radiolabelled D-alanine uptake in wild-type and braG deletion strains, with D-alanine competition for leucine binding to purified BraC.
Type: transport assay
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