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 interact ion with SecA, a translocation ATPase. To examine whether or not the h ydrophobic region of signal peptides also interacts with SecA, we cons tructed model preproteins, proOmpF-Lpps, possessing no positively char ged 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 p ositively charged residues were not required for the translocation of preproteins across the cytoplasmic membrane of Escherichia call both i n vitro and in vivo. Chemical cross-linking between SecA and preprotei ns possessing no positively charged residues at the amino-terminus was observed only in the presence of liposomes containing acidic phosphol ipids. The degree of cross-linking increased as the length of the hydr ophobic stretch increased irrespective of whether positively charged r esidues were present or not. A preprotein possessing no positively cha rged residues at the amino-terminus, which is competent in the presenc e 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 doma ins are involved in the interaction with SecA in the initial stage of translocation in addition to their possible roles in transmembrane mov ement of preproteins.