braD (PA1073, P21627) notes

Identity

UniProt P21627 (Swiss-Prot, PE 3), 307 aa, gene braD, locus PA1073,
P. aeruginosa PAO1. Fetched accession matches the worklist entry (rank 30).
Polytopic inner-membrane permease, with ten predicted TM helices
(UniProt FT TRANSMEM). Pfam PF02653 BPD_transp_2; CDD cd06582
TM_PBP1_LivH_like; PANTHER PTHR11795:SF371 (LIVH). UniProt places it in the
LivHM subfamily. TCDB 3.A.1.4.8.
One of the two permease subunits (BraD, BraE) of LIV-I, the
braC-braD-braE-braF-braG ABC importer (PA1074-PA1070).

Literature

Cloning: BraD is 67% identical to E. coli LivH and is a very hydrophobic
membrane protein
PMID:2120183.
Complementation shows braD is required for LIV-I
PMID:2120184.
Reconstitution: BraDEFG form a membrane complex. With BraC, the complex
transports Leu, the branched-chain amino acids, Ala and Thr in an
ATP-dependent way
PMID:1400443
PMID:1400443.
LIV-I substrate range in cells
PMID:113383.
Johnson et al. 2008 Biolog screen (PMID:18833300, full text; PathoGenesis
Mini-Tn5 mutant, strain PAK). PA1073 grows less well (+/-) on D-Ala (C) and
L-Ile (C), and does not grow on L-Ala (C) (Table 2). No valine phenotype was
recorded for PA1073
PMID:18833300.
The insertion in braD is in the middle of the operon and is likely polar on
braE-braG. The phenotype is therefore an operon phenotype, which is harmless
here because every bra gene is needed.

Interpretation

Same as for braF (see genes/PSEAE/braF/braF-notes.md). L-Leu, L-Ile, L-Val,
L-Ala (and L-Thr) import by BraCDEFG is directly established. D-Ala import is
plausible but rests only on growth phenotypes. Phenylalanine comes from E. coli
LivH and has no P. aeruginosa support.

PseudoCAP IBA-seed assessment

PTN000765102 is the LivH permease node: 18 leaves in 16 taxa (E. coli,
Salmonella and Yersinia LivH, Synechocystis LivH, BraD, PA4912, Geobacter,
Deinococcus (two), Thermotoga, Chloroflexus, Dictyoglomus, Clostridium,
Fusobacterium, Streptococcus, Streptomyces, Gloeobacter, Bradyrhizobium). It
carries E. coli LivH (P0AEX7)-seeded IBDs for L-leucine import, L-valine
transport, and L-Leu/Ile/Val/Phe transporter activity. PTN000209860 is a much
broader permease node (24+ leaves, including archaea (Sulfolobus, Pyrobaculum),
plants (Ricinus, tomato) and the urea permease UrtB), seeded by E. coli LivH
plus the two PseudoCAP rows.

PTN node	node term	n_seeds	leaf IBAs	seed annotation (term, evidence, PMID)	verdict	rationale
PTN000209860	GO:0055085 transmembrane transport	3 (P0AEX7, P21627, Q9HUQ1)	50 (15 taxa)	GO:0042941 and GO:1903806, IMP, PMID:18833300 (PseudoCAP)	SOUND	The generic term suits a broad permease clade that includes urea and other HAAT substrates. It is independently supported by the E. coli LivH seed and by BraDEFG reconstitution.
PTN000765102	GO:1903806 L-isoleucine import across plasma membrane	1 (sole)	18 (16 taxa)	GO:1903806, IMP, PMID:18833300 (PseudoCAP)	SOUND	The growth phenotype is corroborated by LIV-I uptake and BraC binding of Ile (PMID:113383, PMID:6767701). It matches the E. coli-seeded L-isoleucine transporter activity and L-leucine import IBDs at the same node.
PTN000765102	GO:0015808 L-alanine transport	1 (sole)	18 (16 taxa)	GO:0015808, IMP, PMID:18833300 (PseudoCAP)	SOUND	The PA1073 mutant fails to grow on L-Ala, and BraCDEFG transports Ala in proteoliposomes. Correct for P. aeruginosa, but Ala is a secondary substrate set by BraC, and no non-P. aeruginosa seed supports it clade-wide. GO:1904273 L-alanine import across plasma membrane would be more precise.
PTN000765102	GO:0042941 D-alanine transmembrane transport	2 (P21627, Q9HUQ1)	18 (16 taxa)	GO:0042941, IMP, PMID:18833300 (PseudoCAP)	INDIRECT-PHENOTYPE	Both seeds are partial D-Ala growth defects from one Biolog screen. No D-Ala uptake or binding has been shown for LIV-I. Plausible but not established.

No source defects were filed. The L-Ile and L-Ala PseudoCAP rows are true of
LIV-I, and the D-Ala rows are left UNDECIDED, with a suggested question.

Decisions summary

ACCEPT: plasma membrane (IBA, IEA); L-valine, L-isoleucine and L-leucine
transporter activity (IBA); L-leucine import (IBA); L-isoleucine import
(IBA, IMP).
MODIFY: L-alanine transport (IBA, IMP) -> L-alanine import across plasma
membrane; L-valine transmembrane transport (IBA) -> L-valine import across
plasma membrane; membrane (IEA) -> plasma membrane; transmembrane
transporter activity (IEA) -> ABC-type amino acid transporter activity;
transmembrane transport (IEA) -> the four import terms.
MARK_AS_OVER_ANNOTATED: L-phenylalanine transmembrane transporter activity
(IBA), phenylalanine transport (IBA).
UNDECIDED: D-alanine transmembrane transport (IBA, IMP).
NEW: part_of the substrate-binding-subunit-containing ABC transporter
complex (IDA, PMID:1400443).