A peptide corresponding to an export-defective mutant OmpA signal sequence with asparagine in the hydrophobic core is unable to insert into model membranes.
We have examined the comparative membrane interaction properties of synthetic peptides corresponding to the wild-type and an export-defective, mutated signal sequence from the Escherichia coli outer membrane protein, OmpA. As part of a collaborative study of the effects of various alterations on the function of the OmpA signal sequence and the biophysical properties of the corresponding synthetic peptides, we incorporated the small, neutral polar residue, asparagine, into the hydrophobic core in place of Ile-8. This seemingly minor perturbation to the signal sequence caused a complete block of export in vivo (J. Goldstein, S. Lehnhardt, and M. Inouye, following paper). We now explore in detail the difference in the properties of the wild-type and the Ile-8----Asn OmpA signal peptides. The fluorescent residue Trp was substituted in both peptides in place of the wild-type Phe at position 15. This mutation is silent phenotypically and provides a superb probe of membrane interaction. We find that the Asn substitution leaves the conformational properties of the signal sequence essentially unchanged, but prevents any significant interaction of the peptide with a lipid bilayer. Asparagines are very underrepresented among known signal sequences. We believe this low frequency to be due to the lowering of mean residue hydrophobicity caused by incorporation of Asn and the consequent reduced ability to bind and insert into membranes.
Hoyt
D W
DW
Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas 75235-9041.
Gierasch
L M
LM
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D002942:Circular Dichroism; D004926:Escherichia coli / Q000378:metabolism*; D005454:Fluorescence Polarization; D008051:Lipid Bilayers; D009154:Mutation; D021382:Protein Sorting Signals / Q000235:genetics* / Q000378:metabolism; D014364:Tryptophan
D001425:Bacterial Outer Membrane Proteins; D008051:Lipid Bilayers; D021382:Protein Sorting Signals; D014364:Tryptophan