Twin arginine translocation (Tat)-dependent export in the apparent absence of TatABC or TatA complexes using modified Escherichia coli TatA subunits that substitute for TatB.
The twin arginine translocation pathway exports folded proteins across the cytoplasmic membrane of many bacteria. In Escherichia coli and other Gram-negative bacteria, TatA, TatB, and TatC are all essential for efficient translocation, and current models suggest that separate TatABC and TatA complexes coalesce at the point of translocation. However, other microbes appear only to possess tatA and tatC genes. In Escherichia coli, virtually no translocation is observed when only TatA and TatC are present, but several mutations at the extreme N terminus of TatA were shown to support translocation. Here we show that these apparently bifunctional mutant TatA variants can function as typical TatA components because translocation is observed when they are co-expressed with TatBC, and they assemble into large, heterogeneous complexes that resemble wild type TatA complexes. However, cells expressing TatC plus the mutant TatA variants do not contain complexes that resemble the expected 370-kDa TatABC complex, clearly indicating that the mutant TatA forms cannot assemble efficiently, or stably, into this complex. The simultaneous expression of wild type TatA furthermore blocks translocation activity, suggesting that the mutant TatA forms preferentially bind to other TatA molecules rather than TatC. Surprisingly, we observe translocation in the absence of detectable free TatA, when translational fusions of the mutant TatAs with TatC are expressed. Transport can thus proceed in the simultaneous absence of TatABC and TatA complexes at detectable levels, and we conclude that the active translocon may be formed from dynamic twin arginine translocation complexes, one or more of which may await characterization.
Barrett
Claire M L
CM
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, United Kingdom.
Freudl
Roland
R
Robinson
Colin
C
D026923:Amino Acid Transport Systems, Basic / Q000235:genetics / Q000378:metabolism*; D001120:Arginine / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism*; D009154:Mutation
D026923:Amino Acid Transport Systems, Basic; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D046912:Multiprotein Complexes; C499697:TatA protein, E coli; C473116:TatB protein, E coli; C461379:TatC protein, E coli; D001120:Arginine