Twin-arginine-dependent translocation of folded proteins

Abstract

Twin-arginine translocation (Tat) denotes a protein transport pathway in bacteria, archaea and plant chloroplasts, which is specific for precursor proteins harbouring a characteristic twin-arginine pair in their signal sequences. Many Tat substrates receive cofactors and fold prior to translocation. For a subset of them, proofreading chaperones coordinate maturation and membrane-targeting. Tat translocases comprise two kinds of membrane proteins, a hexahelical TatC-type protein and one or two members of the single-spanning TatA protein family, called TatA and TatB. TatC- and TatA-type proteins form homo- and hetero-oligomeric complexes. The subunits of TatABC translocases are predominantly recovered from two separate complexes, a TatBC complex that might contain some TatA, and a homomeric TatA complex. TatB and TatC coordinately recognize twin-arginine signal peptides and accommodate them in membrane-embedded binding pockets. Advanced binding of the signal sequence to the Tat translocase requires the proton-motive force (PMF) across the membranes and might involve a first recruitment of TatA. When targeted in this manner, folded twin-arginine precursors induce homo-oligomerization of TatB and TatA. Ultimately, this leads to the formation of a transmembrane protein conduit that possibly consists of a pore-like TatA structure. The translocation step again is dependent on the PMF.

Figure 1.

Tat signal sequences. Shown are the general composition of (top) Tat signal sequences and the amino acid sequences of three natural Tat precursor proteins of E. coli. SufI and TorA are the most common Tat substrates used for investigations of the Tat pathway and CueO is a copper oxidase, the RR-signal peptide of which comes closest to the consensus motif.

Figure 2.

The Tat components. (a) The known representatives of the TatA-protein family, TatA, TatB, TatE are depicted according to a recent nuclear magnetic resonance structure solved for an N-terminal fragment of the B. subtilis paralogue TatA_d [36]. (b) Sequence alignment of the conserved N-terminal fragments of TatA, TatE and TatB from E. coli and TatA_d and TatA_y from B. subtilis. Numbering refers to the residues of E. coli TatA. The transmembrane and amphipathic helices and the intervening hinge region are indicated according to references [–39]. (c) Predicted structure of TatC of E. coli with the approximate positions of residues that influence its activity and the putative substrate binding site highlighted in red. (d) The major Tat complexes recovered from TatABC translocases of Gram-negative bacteria and thylakoids of plant chloroplast. TatA forms homo-oligomeric complexes and is represented here by a tetramer. TatB and TatC form stable complexes independently of TatA. When purified from Gram-negative bacteria, not from chloroplasts, TatBC complexes often contain TatA.

Figure 3.

Model of a substrate-induced formation of a functional Tat translocase. (a) Step 1: superficial binding of a Tat signal sequence to the Tat translocase involves the RR-consensus motif and cytosolic parts of TatC. The coordinate recognition of a Tat signal sequence by TatC and TatB leads to its hairpin-like insertion into the plane of the membrane. Step 2: TatB homo-oligomerizes and encapsulates the folded mature domain of the substrate. Step 3: a proton motive-force-dependent step brings monomeric TatA in close contact to the signal sequence. AxA denotes the C-terminal end of the signal sequence. Step 3 is also shown as top view from the periplasmic side of the membrane. (b) More TatA subunits are recruited from a pool of monomeric TatA or more likely in tetrameric units. Processive oligomerization leads to the pavement of a transmembrane path. This might comprise oligomeric TatA pores, hetero-oligomeric TatA(BC) pores as drawn here or some other undefined TatA structure.