The hydrophobic region of signal peptides is involved in the interaction with membrane-bound SecA.

The positive charges of signal peptides are important for the interaction with SecA, a translocation ATPase. To examine whether or not the hydrophobic region of signal peptides also interacts with SecA, we constructed model preproteins, proOmpF-Lpps, possessing no positively charged amino acid residues at the amino-terminus and different numbers of alanine/leucine residues in the hydrophobic region of signal peptides. When the hydrophobic stretch was sufficiently long, amino-terminal positively charged residues were not required for the translocation of preproteins across the cytoplasmic membrane of Escherichia coli both in vitro and in vivo. Chemical cross-linking between SecA and preproteins possessing no positively charged residues at the amino-terminus was observed only in the presence of liposomes containing acidic phospholipids. The degree of cross-linking increased as the length of the hydrophobic stretch increased irrespective of whether positively charged residues were present or not. A preprotein possessing no positively charged residues at the amino-terminus, which is competent in the presence of liposomes, competitively inhibited the cross-linking of wild-type proOmpF-Lpp with SecA under the same conditions. It is concluded that both the amino-terminal positive charges and central hydrophobic domains are involved in the interaction with SecA in the initial stage of translocation in addition to their possible roles in transmembrane movement of preproteins.