Affinity of TatCd for TatAd Elucidates Its Receptor Function in the Bacillus subtilis Twin Arginine Translocation (Tat) Translocase System

Abstract

Twin arginine translocation (Tat) systems catalyze the transport of folded proteins across the bacterial cytosolic membrane or the chloroplast thylakoid membrane. In the Tat systems of Escherichia coli and many other species TatA-, TatB-, and TatClike proteins have been identified as essential translocase components. In contrast, the Bacillus subtilis phosphodiesterase PhoD-specific system consists only of a pair of TatA In bacteria, most of the exported proteins cross the cytosolic membrane through the Sec-dependent translocase in an unfolded conformation (1-3). A subset of proteins is transported via the twin arginine translocation (Tat) 2 pathway. This alternative route accepts folded proteins or protein domains as substrates (4 -6). Initially, the Tat pathway was discovered in chloroplasts as a pathway that operates independently of soluble factors and nucleoside triphosphates and is exclusively energized by the proton gradient across the membrane (7-9). In vitro translocation systems using Escherichia coli components demonstrated that also in bacteria this transport system is energized exclusively by the transmembrane proton electrochemical gradient (10,11). Substrates destined for export by the Tat system are synthesized as preproteins with a signal peptide containing an almost invariant twin arginine sequence motif in the N-region of the otherwise canonically structured signal sequence (12).In E. coli TatA, TatB, and TatC proteins have been demonstrated to be essential for Tat-dependent protein transport (13-16). The TatA homologous protein TatE has been proven to be functionally redundant (17). In chloroplasts structural and functional counterparts Tha4 (homologous to TatA; Ref. 18), Hcf106 (homologous to TatB; Refs. 16, 19), and cpTatC (homologous to TatC; Ref. 20) have been identified. The sequence-related proteins TatA and TatB are anchored in the cytoplasmic membrane via an amino-proximal ␣-helical domain (13). TatCs are a family of proteins with six calculated transmembrane-spanning domains with N and C termini exposed to the cytoplasmic or stromal side of the membrane (4). Gouffi et al. (21) recently proposed function-linked changes of TatA and TatC topologies for the mechanism of folded protein translocation in E. coli. These changes involve a TatA, the C terminus of which shuttles between the cytosol and the periplasmic space, and a TatC, the predicted fourth and fifth transmembrane helices of which shuttle in the periplasmic space. They speculated that this topology of TatC might reflect an operational state of TatC that changes during the protein translocation process (21). The current proposal for the action of Tat transport system of E. coli and plant thylakoids involves the initial binding of substrates to the TatB-TatC high molecular weight complex mediated via a direct contact of the double arginine signal peptide with TatC (20,22). The signal peptide binding triggers association with TatA to form the active translocation channel driven by the transmembrane proton electrochemical gradien...

The Tat pathway in Streptomyces lividans: interaction of Tat subunits and their role in translocation

“…Two possibilities are considered: (1) TatA and TatB interact with the protein and recognize either the same or a different region of the signal peptide; (2) either TatA or TatB interacts in vivo with the Tat substrate. Very recently, it was reported that B. subtilis TatA molecules are organized as cylindermicelle-like structures with a 120 Å hydrophobic centre, which could accommodate an a-helical domain of a signal peptide (Westermann et al, 2006). As we have observed hetero-oligomerization of TatA and TatB in the cytoplasm of S. lividans, it is very likely that they are organized in similar structures.…”

“…The function of TatA as being involved in pore formation and mediating transport of its Tat substrate across the cytoplasmic membrane seems to be very conserved in S. lividans, E. coli and B. subtilis. In E. coli and B. subtilis, a spectrum of TatA complexes forming ringlike structures in the membrane (Gohlke et al, 2005;Westermann et al, 2006) has been reported as well as the colocalization of Tat precursors with membrane-integrated TatA (Alami et al, 2003;Westermann et al, 2006). In addition, cross-complementation tests recently revealed that Streptomyces coelicolor TatA protein supports a high level of Tat activity in an E. coli double tatA/E mutant.…”

Recombinant Expression of tatABC and tatAC Results in the Formation of Interacting Cytoplasmic TatA Tubes in Escherichia coli

“…In addition, because of the co-fractionation of TatA with membranes, which depended on the hydrophobic N-terminal region of the protein, it has been assumed that TatA must be an exclusively membrane-bound component of the Tat system (35). However, electron microscopy of TatA d from Bacillus subtilis revealed that a significant portion of this TatA was present in the cytoplasm where it was able to bind the Tat substrate PhoD (8,26). The TatA d , which interacted with membranes, had substrate bound to it (9).…”

Subcellular Localization of TatAd of Bacillus subtilis Depends on the Presence of TatCd or TatCy

“…Despite the fact that it contains an N-terminal transmembrane segment (17), TatAd was also found in the cytosol, where it appears to interact with its substrate, pre-PhoD, via the signal sequence (24). TatCd was proposed to act as a receptor for the anchoring at and subsequent incorporation into the membrane of this TatAd-PhoD complex (28).…”