The conformation of a signal peptide bound by Escherichia coli preprotein translocase SecA.
To understand the structural nature of signal sequence recognition by the preprotein translocase SecA, we have characterized the interactions of a signal peptide corresponding to a LamB signal sequence (modified to enhance aqueous solubility) with SecA by NMR methods. One-dimensional NMR studies showed that the signal peptide binds SecA with a moderately fast exchange rate (Kd approximately 10(-5) m). The line-broadening effects observed from one-dimensional and two-dimensional NMR spectra indicated that the binding mode does not equally immobilize all segments of this peptide. The positively charged arginine residues of the n-region and the hydrophobic residues of the h-region had less mobility than the polar residues of the c-region in the SecA-bound state, suggesting that this peptide has both electrostatic and hydrophobic interactions with the binding pocket of SecA. Transferred nuclear Overhauser experiments revealed that the h-region and part of the c-region of the signal peptide form an alpha-helical conformation upon binding to SecA. One side of the hydrophobic core of the helical h-region appeared to be more strongly bound in the binding pocket, whereas the extreme C terminus of the peptide was not intimately involved. These results argue that the positive charges at the n-region and the hydrophobic helical h-region are the selective features for recognition of signal sequences by SecA and that the signal peptide-binding site on SecA is not fully buried within its structure.
Chou
Yi-Te
YT
Department of Chemistry, University of Massachusetts, Amherst, Massachusetts 01003-04510, USA.
Gierasch
Lila M
LM
D000251:Adenosine Triphosphatases / Q000737:chemistry*; D000595:Amino Acid Sequence; D001120:Arginine; D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D026901:Membrane Transport Proteins / Q000737:chemistry*; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011522:Protons; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D055672:Static Electricity
D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; D010455:Peptides; D021382:Protein Sorting Signals; D011522:Protons; D000069816:SEC Translocation Channels; D001120:Arginine; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins