The Escherichia coli TatABC system and a Bacillus subtilis TatAC-type system recognise three distinct targeting determinants in twin-arginine signal peptides.
The Tat system transports folded proteins across bacterial and thylakoid membranes. In Gram-negative organisms, it is encoded by tatABC genes and the system recognizes substrates bearing signal peptides with a conserved twin-arginine motif. Most Gram-positive organisms lack a tatB gene, indicating major differences in organisation and/or mechanism. Here, we have characterized the essential targeting determinants that are recognized by a Bacillus subtilis TatAC-type system, TatAdCd. Substitution by lysine of either of the twin-arginine residues in the TorA signal peptide can be tolerated, but the presence of twin-lysine residues blocks export completely. We show that additional determinants can be as important as the twin-arginine motif. Replacement of the -1 serine by alanine, in either the TorA or DmsA signal peptide, almost blocks export by either the B. subtilis TatAdCd or Escherichia coli TatABC systems, firmly establishing the importance of this -1 residue in these signal peptides. Surprisingly, the +2 leucine in the DmsA signal peptide (sequence SRRGLV) appears to play an equally important role and substitution by alanine or phenylalanine blocks export by both the B. subtilis and E. coli systems. These data identify three distinct determinants, whose importance varies depending on the signal peptide in question. The data also show that the B. subtilis TatAdCd and E. coli TatABC systems recognize very similar determinants within their target peptides, and exhibit surprisingly similar responses to mutations within these determinants.
Mendel
Sharon
S
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK.
McCarthy
Andrew
A
Barnett
James P
JP
Eijlander
Robyn T
RT
Nenninger
Anja
A
Kuipers
Oscar P
OP
Robinson
Colin
C
D000409:Alanine / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001120:Arginine / Q000235:genetics*; D001412:Bacillus subtilis / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016384:Consensus Sequence; D004926:Escherichia coli / Q000378:metabolism*; D005798:Genes, Bacterial; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010649:Phenylalanine / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000378:metabolism; D013045:Species Specificity; D013379:Substrate Specificity
D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D012694:Serine; D010649:Phenylalanine; D001120:Arginine; D008239:Lysine; D000409:Alanine