Abstract
Tripartite ATP-dependent periplasmic (TRAP) transporters are widespread in prokaryotes, but absent in eukaryotes, and facilitate the uptake of a wide variety of substrates. TRAP transporters are composed of a substrate binding protein (SBP) and two unequally sized membrane components that exist as either separate proteins or are fused into a single polypeptide. Almost all TRAP SBPs exist as monomeric globular proteins that bind substrate and present it to the membrane component. Here, we describe the discovery and characterisation of a novel TRAP SBP from the TAXI subfamily with a previously unidentified architecture. BP0403 from the human pathogen Bordetella pertussis is a predicted lipoprotein composed of 3 distinct domains; an α/β globular domain with a unique fold, a long helical domain and a C-terminal TAXI SBP domain. Expression and purification of the full-length protein reveals that it forms a stable dimer. Structural modelling of the dimer interface and molecular weight analysis using size exclusion chromatography of the individual domains reveals that the interdomain helical region is solely responsible for dimerization. Differential scanning fluorimetry (DSF) and intrinsic tyrosine fluorescence reveal that BP0403 binds L-glutamate with nanomolar affinity. Unexpectedly, analysis of the genome context of BP0403 reveals the complete absence of characteristic genes for TRAP membrane components but co-localisation and translational coupling with gltS, encoding a Na+/glutamate symporter. In other bacteria, we identified fused BP0403-GltS homologues, strongly suggesting that this constitutes a completely novel SBP-dependent secondary active transporter. Structural comparisons suggest GltS operates by an elevator-type mechanism, like TRAP transporters; the association of an SBP with this class of secondary transporter is an emerging theme.
Competing Interest Statement
The authors have declared no competing interest.
