Conformational and membrane-binding properties of a signal sequence are largely unaltered by its adjacent mature region

Abstract

We have synthesized a peptide corresponding to the 25-residue signal sequence plus the first 28 residues of the Escherichia coli outer membrane protein LamB in order to explore the properties of a signal sequence in the presence of the N-terminal region of its passenger. In the last few years, there have been several observations of differing efficiencies of export when signal sequences are attached to different passenger proteins or when the first part of a passenger protein undergoes mutation. In the LamB case, gene fusions with lacZ have shown that the signal sequence plus the first 28 residues of mature LamB are necessary to direct beta-galactosidase into the export pathway [Rasmussen, B. A. & Silhavy, T. J. (1987) Genes Dev. 1, 185-196]. The origin of these observations and whether there is an influence of the mature region on the properties of the signal sequence have not been known. We find that the conformational and membrane-binding properties of the LamB signal sequence manifest in a 25-residue peptide are essentially unaltered in the context of the 53-residue peptide corresponding to this signal sequence plus the first 28 residues of the mature LamB protein. CD spectra show that the signal peptide and passenger domains are conformationally independent of each other in micelle or bilayer environments. Furthermore, the signal sequence leads to the spontaneous association of the 53-residue peptide with a lipid bilayer; alone, the mature domain does not interact with lipid bilayers. Fluorescence results show that the mode of interaction of the signal peptide with a bilayer is essentially unaltered by the presence of its mature region. This lack of influence of the mature domain on the behavior of the signal sequence is unexpected for juxtaposed polypeptides of comparable length and may be of physiological importance: N-terminal regions of secreted proteins may be selected to be passive, by comparison with their cognate signal sequences, which themselves must engage the export apparatus and actively interact with its components.