EFFECT OF CHARGED RESIDUE SUBSTITUTIONS ON THE THERMODYNAMICS OF SIGNAL PEPTIDE-LIPID INTERACTIONS FOR THE ESCHERICHIA-COLI LAMB SIGNAL SEQUENCE

We have used tryptophan fluorescence spectroscopy to characterize the binding affinities of an Escherichia coli LamB signal peptide family f or lipid vesicles. These peptides harbor charged residue substitutions in the hydrophobic core region. Titrations of peptides with vesicles composed of almitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine and 1-p almitoyl-2-oleoyl-sn-3-phosphoglycerol (65:35 mol%), in conjunction wi th evaluation of peptide dissociation rates from these vesicles, were used to determine binding parameters quantitatively. We find that unde r low ionic strength conditions, point mutations introducing negativel y charged aspartate residues substantially reduce peptide affinity rel ative to the wild-type peptide. However, the difference between wild-t ype and mutant peptide affinities was much lower under approximately p hysiological ionic strength. In addition, the lipid affinities of mode l surface-binding and transmembrane peptides were determined. These co mparative studies with signal and model peptides permitted semi-quanti tative deconvolution of signal peptide binding into electrostatic and hydrophobic components. We find that both interactions contribute sign ificantly to binding, although the theoretically available hydrophobic free energy is largely offset by unfavorable polar-group effects. The implications of these results for understanding the potential roles o f the signal sequence in protein translocation are discussed.