Interaction of mutant alkaline phosphatase precursors with membrane phospholipids in vivo and in vitro

Positively charged amino acid residues in the N-terminal domain of the sign al peptides of secreted proteins are thought to interact with negatively ch arged anionic phospholipids during the initiation of secretion. To test thi s hypothesis, substitutions of the uncharged Ala or the negatively charged Glu residue for the positively charged Lys-20 of the N-terminus of the sign al peptide of Escherichia coli alkaline phosphatase were introduced using a modified method of oligonucleotide-directed mutagenesis. We found that Lys -20 is involved in the interaction of the signal peptide with anionic phosp holipids in viva and effects the efficiency of insertion of the signal pept ide of isolated precursor into model phospholipid membranes in vitro. We al so show that the efficiency of signal peptide insertion into the lipid bila yer depends on the fluidity of the bilayer.