Abstract

A synthetic peptide corresponding to the signal sequence of wild type Escherichia coli lambda-receptor protein (LamB) inhibits in vitro translocation of precursors of both alkaline phosphatase and outer membrane protein A into E. coli membrane vesicles (half-maximal inhibition at 1-2 microM). By contrast, the inhibitory effect was nearly absent in a synthetic peptide corresponding to the signal sequence from a mutant strain that harbors a deletion mutation in the LamB signal region and displays an export-defective phenotype for this protein in vivo. Two peptides derived from pseudorevertant strains that arose from the deletion mutant and exported LamB in vivo were found to inhibit in vitro translocation with effectiveness that correlated with their in vivo export ability. Controls indicated that these synthetic signal peptides did not disrupt the E. coli membrane vesicles. These results can be interpreted to indicate that the presequences of exported proteins interact specifically with a receptor either in the E. coli inner membrane or in the cytoplasmic fraction. However, biophysical data for the family of signal peptides studied here reveal that they will spontaneously insert into a lipid membrane at concentrations comparable to those that cause inhibition. Hence, an indirect effect mediated by the lipid bilayer of the membrane must be considered.

Highlights

  • Vesicles Is Inhibited by Functional of proteins is thesignal or presequence that occurs as an N

  • A synthetic peptide corresponding to the signal sequence of wild type Escherichia coliX-receptorprotein (LamB)inhibits in vitrotranslocation of precursors of both alkaline phosphataseand outer membraneprotein A into E. coli membrane vesicles

  • The inhibitory effect was nearly absent in a synthetic peptide corresponding to the signal sequence from a mutant strain that harbors a deletion mutation in the LamB signal region and displays an export-defective phenotype for this protein in vivo

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Summary

Synthetic SignalPeptides*

Terminalextensiononthematureprotein.Recentresults suggest that anenergy requirement, andmore an (Received for publication, October 10, 1986) ATPrequirement [4,5,6], may be quite general [7,8,9,10,11]. Controls indicated that sequence was derived.A“consensus” eukaryotic signal sethese synthetic signal peptides did not disrupt the E. quence designed by Austen andco-workers [20] inhibited coli membrane vesicles. These results can beinter- a range of normally translocated proteins. Quencesare derived fromthe wild type X-phage receptor (LamB)’ signal peptide,an export-defective mutantstrain that accumulates precursor to theX-receptor as a result of a 4-residue deletion in its signal sequence, and two pseudorevertant strains that arose from thdiesletion mutant by point mutations [26] (Fig. 1).We found previously that theabilities f To whom correspondence should he addressed.

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RESULTS
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