2.A.56 The Tripartite ATP-independent Periplasmic Transporter (TRAP-T) Family
TRAP-T family permeases generally consist of three components, and these systems have so far been found in Gram-negative bacteria, Gram-positive bacteria and archaea. Several members of the family have been both sequenced and functionally characterized. The first system to be characterized was the DctPQM system of Rhodobacter capsulatus (Forward et al., 1997), and it is the prototype for the TRAP-T family (Kelly and Thomas, 2001; Rabus et al., 1999).
DctP is a periplasmic dicarboxylate (malate, fumarate, succinate) binding receptor that is biochemically well-characterized. The 3-dimensional structure of a homologue, SiaTP (TC #2.A.56.1.3) has been solved (Muller et al., 2006). DctQ is an integral cytoplasmic membrane protein (25 kDa) with 4 putative transmembrane α-helical spanners (TMSs). DctM is a second integral cytoplasmic membrane protein (50 kDa) with 12 putative TMSs. These three proteins have been shown to be both necessary and sufficient for the proton motive force-dependent uptake of dicarboxylates into R. capsulatus. An involvement of ATP in transport energization was excluded. The substrate-binding protein, SiaP, imposes directionality on an electrochemical sodium gradient-driven TRAP transporter, SiaPQM (Mulligan et al., 2009).
In several TRAP-T systems, fused Q-M-type proteins instead of two separate Q- and M-type proteins are found, while in others, Q-P-type fusion proteins are found. The operon encoding the Synechocystis system includes a protein homologous to the glutamine binding protein, and biochemical evidence has suggested that a glutamate transporter from Rhodobacter sphaeroides is a periplasmic binding protein-dependent, pmf-dependent secondary carrier (Jacobs et al., 1996). Homologous systems in Halomonas elongata and Rhodobacter spheroides take up ectoine/hydroxyectoine and taurine, respectively (Bruggemann et al., 2004; Grammann et al., 2002). The DctP dicarboxylate receptor is homologous to both the YiaO monocarboxylate receptor and the TeaA ectoine receptor. Thus, the TRAP-T family of permeases may be involved in the uptake of widely divergent compounds, mostly carboxylate derivatives (Kelly and Thomas, 2001; Thomas et al., 2006; Mulligan et al., 2007).
The crystal structure of SiaP (the receptor for SiaTP; TC #2.A.56.1.3) reveals an overall topology similar to ATP binding cassette receptors, which is not apparent from the sequence, demonstrating that primary and secondary transporters can share a common structural component (Müller et al., 2006). The structure of SiaP in the presence of the sialic acid analogue 2,3-didehydro-2-deoxy-N-acetylneuraminic acid reveals the ligand bound in a deep cavity with its carboxylate group forming a salt bridge with a highly conserved Arg residue. Sialic acid binding, which obeys simple bimolecular association kinetics, is accompanied by domain closure about a hinge region and the kinking of an α-helix hinge component. The structure provides insight into the evolution, mechanism, and substrate specificity of TRAP-transporters (Müller et al., 2006).
The solute binding receptor, DctP, has a structure comprised of two domains connected by a hinge that closes upon substrate binding, similar to those in ABC uptake porters. Substrate binding is mediated through a conserved and specific arginine/carboxylate interaction in the receptor. Mulligan et al. (2011) have reviewed the expanding repertoire of substrates and physiological roles for experimentally characterized TRAP transporters in bacteria and discuss mechanistic aspects. TRAP transporters are high-affinity, Na+-dependent unidirectional secondary transporters.
A subfamily of TRAP-Ts [tetratricopeptide repeat-protein associated TRAP transporters (TPATs)] has four components. Three are common to both TRAP-Ts and TPATs. TPATs are distinguished from TRAP-Ts by the presence of a protein called the 'T component'. In Treponema pallidum, this protein (TatT) is a water-soluble trimer whose protomers are each perforated by a pore. Its respective P component (TatP(T)) interacts with TatT. Co-crystal structures of two complexes showed that up to three monomers of TatP(T) can bind to the TatT trimer. A putative ligand-binding cleft of TatP(T) aligns with the pore of TatT, strongly suggesting ligand transfer between T and P(T) (Brautigam et al., 2012).
The generalized transport reaction presumed to be catalyzed by TRAP-T family permeases is:
solute (out) + nH+ (out) → solute (in) + nH+ (in)
Transporter for lignin derived aromatic compounds, TarPQM (Salmon et al. 2013). The purple photosynthetic bacterium Rhodopseudomonas palustris is able to grow photoheterotrophically under anaerobic conditions on a range of phenylpropeneoid lignin monomers, including coumarate, ferulate, caffeate, and cinnamate. TarPQM is encoded at the same locus as CouPSTW (TC# 3.A.1.4.11) and several other genes involved in coumarate metabolism. The periplasmic binding-protein of this system (TarP) binds coumarate, ferulate, caffeate, and cinnamate with nanomolar KD values. Thus, R. palustris uses two redundant but energetically distinct primary and secondary transporters that both employ high-affinity periplasmic binding-proteins to maximize the uptake of lignin-derived aromatic substrates from the environment (Salmon et al. 2013).
TarPQM of Rhodopseudomonas palustris
TarP (R; 336 aas)
TarQ (small M; 217 aas)
TarM (large M; 435 aas)
Tripartite high affinity ectoine/hydroxyectoine uptake system (Grammann et al., 2002). Deletion leads to increased rates of ectoine excretion (Hobmeier et al. 2022). In the absence of the substrate-binding protein, TeaA, an overexpression of both subunits TeaBC facilitated a three-fold increased excretion rate of ectoine export. Individually, the large subunit TeaC showed an approximately five times higher extracellular ectoine concentration per dry weight compared to TeaBC shortly after its expression was induced. This led to the possibility that only the large subunit, TeaC, is required for channel function (Hobmeier et al. 2022).
TeaABC ectoine transporter of Halomonas elongata
TeaB (M, 4 TMS)
TeaC (M, 12 TMS)
2-Oxoglutarate, 2OG (α-ketoglutarate, αKG) uptake porter of 677 aas and 20 TMSs (Large + small subunits fused) plus a periplasmic solute binding protein of 318 aas and 1 N-terminal TMS.
αKG uptake porter of Shewanella oneidensis
DctM4Q4P4 three component TRAP-T transporter that may take up phenylacetate and phenylpyruvate (Dörries et al. 2016).
DctM4Q4P4 of Desulfococcus multivorans
Three component TRAP-T transporter, DctM9Q9P9; may take up phenylacetate and phenylalanine (Dörries et al. 2016).
TRAP-T uptake system of Desulfobacula toluolica Tol2
DctM9, 430 aas and 14 TMSs; 90% identical to DctM4 (TC# 2.A.56.1.14)
DctQ9, 160 aas and 4 TMSs; 75% identical to DctQ4 (TC# 2.A.56.1.14)
DctP9, 358 aas and 1 TMS; 79% identical to DctP4 (TC# 2.A.56.1.14)
Uncharacterized TRAP-T family with DctM, DctQ and DctP; may transport dicarboxylic acids (by similarity).
TRAP-T family system of Gammaproteobacteria bacterium (marine metagenome)
DctM, 433 aas, 10 TMSs, MBI80045
DctQ, 168 aas and 4 TMSs, MBI80046
DctP, 377 aas and 1 N-terminal TMS, MBI80047
2-oxoglutarate transporter with two components, a 20 - 22 TMS integral membrane protein of 674 aas and a solute-binding receptor with 317 aas and one N-terminal TMS. The former protein is 81% identical to the large membrane protein with TC# 2.A.56.1.12, and the latter is 74% identical to the binding protein constituent of TC# 2.A.56.1.12. The former two membrane protein homologues seem to have the same topologies with 20 - 22 TMSs.
2-oxoglutarate uptake TRAP transporter of Pseudomonas stutzeri
TRAP transporter with one membrane constituent (743 aas and 22 TMSs) and one receptor (330 aas and 1 N-terminal TMS). May transport dicarboxyic acids: 2-oxoglutarate, fumarate, L-malate and succinate. The membrane constituent is 37% identical to that in TC# 2.A.56.1.17, and the receptor is 21% identical to the receptor in TC# 2.A.56.1.17.
TRAP transporter of Dinoroseobacter shibae
The putative outer membrane anion-selective porin, TsaT, of 338 aas and probably 1 N-terminal TMS (Mampel et al. 2004). Although it was reported to be an outer membrane porin, it is homologous to periplasmic binding receptors of the TRAP-T family. It previously had TC# 9.A.56.1.1.
TsaT of Comamonas testosteroni (Pseudomonas testosteroni) (Q8KR68)
Na+-dependent (smf-driven) sialic acid (N-acetyl neuraminic acid) transporter, SiaTP (Allen et al., 2005; Severi et al., 2005; Johnston et al., 2008). SiaT is also called SiaQM (Mulligan et al., 2009). Also transports the related sialic acids, N-glycolylneuraminic acid (Neu5Gc) and 3-keto-3-deoxy-D-glycero-D-galactonononic acid (KDN) (Hopkins et al. 2013).
SiaTP of Haemophilus influenzae
SiaT (a fusion protein equivalent to both DctM and DctQ) (616 aas; 16 TMSs) (P44543)
SiaP (R) (P44542)
Putative tripartite taurine uptake system, TauKLM (Bruggemann et al., 2004; Denger et al., 2006)
TauKLM of Rhodobacter sphaeroides
TauK (Rsph2615) (R) (Q3IVI6)
TauL (Rsph2614) (M, 4 TMS) (Q3IVI5)
TauM (Rsph2613) (M, 12 TMS) (Q3IVI4)
The putative rhamnogalacturonide transporter (Rodionov et al. 2004)
RhiABC of Salmonella typhimurium
RhiA (R) (P43020)
RhiB (M, 4 TMSs) (Q8ZKR9)
RhiC (M, 12 TMSs) (Q8ZKS0)
The Na+-dependent sialic acid uptake porter, SiaPQM. SiaQ and SiaM form a 1:1 stoichiometric complex (Mulligan et al., 2012). The structure of a Vibrio ortholog has been determined by cryoEM (Peter et al. 2022). The protein complex is composed of 16 TMSs in SiaQ (4 TMSs) and SiaM (12 TMSs) that are structurally related to multimeric elevator-type transporters. The idiosyncratic Q-domain of TRAP transporters enables the formation of a monomeric elevator architecture. A model of the tripartite PQM complex is experimentally validated and reveals the coupling of the substrate-binding P protein to the transporter domains. Peter et al. 2022 studied the formation of the tripartite complex and investigated the impact of interface mutants.
SiaPQM of Vibrio cholerae
SiaP (R) (Q9KR64)
SiaQ (M, 4TMSs) (B9TSN0)
SiaM (M, 12 TMSs) (B9TSM9)
The malonate uptake transporter, MatPQM. Regulated by the GtrA transcriptional activator (Chen et al. 2010). MatM is fused in a single protein C-terminal to MatA (malonyl-CoA decarboxylase).
MatPQM of Sinorhizobium meliloti
MatP (R) (Q930W1)
MatQ (M, 4TMSs) (Q930W2)
MatM (M, 12TMSs) (Q930W3)
Sialic acid uptake transporter, DctMPQ
DctMPQ of E. coli
The possible disulfide 3,3'-dithiodipropionic acid (DTDP) tripartite transporter, DctMPQ (Wübbeler et al. 2014). More probably takes up an array of oxidized sugar onic acids, D-gluconate, D-galactonate, L-arabonate, D-fuconate and D-xylonate. The sugars are oxidized by a broad-range, membrane bound sogar oxidase. The acids that have been studied kineticall have Kms between 8 and 15 μM (Meinert et al. 2017; ).
DTDP transporter of Advenella mimigardefordensis strain DPN7
DctM (M, large)
DctQ (M, small)
TRAP transporter for a hydrophobic substrate (3-d structure known; tp0958 has 18-20 TMSs) (Deka et al., 2012). The substrate could be a lipoprotein, tp0956 (O83922) which is encoded in the same operon with tp0957 and tp058. This protein differs from all other members of the TRAP-T family in having 19 predicted TMSs with extra TMSs at its N-terminus.
TRAP-T transporter of Treponema pallidum
tp0957 (R) (O83923)
tp0958 (M) (O83924)
The 2-oxo monocarboxylate transporter (Pernil et al., 2010). Transports pyruvate which is inhibited by various 2-ketoacids.
The 2-oxo monocarboxylate transporter of Anabaena (nostoc) sp. strain PCC7120
DctQ (Alr3026) (Q8YSQ8)
DctM (Alr3027) (Q8YSQ7)
DctP (Alr3028) (Q8YSQ6)
The 2-ketomonocarboxylate transporter (presented in order of affinity - 2-oxovalerate [highest affinity, KD=0.1 μM], 2-oxoisovalerate, 2-oxobutyrate, 2-oxoisocaproate, 2-oxo-3-methylvalerate, pyruvate [lowest affinity, KD=3 μM]) (Thomas et al., 2006).
The 2-ketomonocarboxylate transporter of Rhodobacter capsulatus
DctM-2, M-large (D5ATK1)
DctQ-2, M-small (D5ATK0)
DctP-2, Receptor (R) (D5ALT6)