Signal Recognition Particle (SRP)-mediated Targeting and Sec-dependent Translocation of an Extracellular Escherichia coli Protein

Robert Sijbrandi,Malene L Urbanus,Corinne M Ten Hagen-Jongman,Harris D Bernstein,Bauke Oudega,Ben R Otto,Joen Luirink

doi:10.1074/jbc.m211630200

Abstract

Hemoglobin protease (Hbp) is a hemoglobin-degrading protein that is secreted by a human pathogenic Escherichia coli strain via the autotransporter mechanism. Little is known about the earliest steps in autotransporter secretion, i.e. the targeting to and translocation across the inner membrane. Here, we present evidence that Hbp interacts with the signal recognition particle (SRP) and the Sec-translocon early during biogenesis. Furthermore, Hbp requires a functional SRP targeting pathway and Sec-translocon for optimal translocation across the inner membrane. SecB is not required for targeting of Hbp but can compensate to some extent for the lack of SRP. Hbp is synthesized with an unusually long signal peptide that is remarkably conserved among a subset of autotransporters. We propose that these autotransporters preferentially use the co-translational SRP/Sec route to avoid adverse effects of the exposure of their mature domains in the cytoplasm.

Highlights

From the ‡Department of Molecular Microbiology, Vrije Universiteit, De Boelelaan 1087, 1081 HV Amsterdam, The Netherlands and the §Genetics and Biochemistry Branch, NIDDK, National Institutes of Health, Bethesda, Maryland 20892-1810
Hemoglobin protease (Hbp)1 is secreted by a human pathogenic Escherichia coli strain [1] and contributes to the pathogenic synergy between E. coli and Bacteroides fragilis in intraabdominal infections [2]
In the absence of a functional signal recognition particle (SRP) pathway, part of the mistargeted Hbp is rescued by SecB, underscoring the inherent flexibility of protein targeting in E. coli [20]

Summary

The abbreviations used are

Hemoglobin protease; IMP, inner membrane protein; IMV, inverted membrane vesicle; SRP, signal recognition particle; DSS, 2,2-dimethyl-2-silapentanesulfonic acid; TF, trigger factor; TPS, two-partner secretion. Most periplasmic and outer membrane proteins synthesized with a cleavable signal peptide are translocated through the Sec-translocon. Targeting of IMPs to the Sectranslocon is not mediated by SecB but by the signal recognition particle (SRP) and its receptor, FtsY, in a co-translational mechanism that resembles targeting to the Sec complex in the endoplasmic reticulum [8]. We provide evidence that the long signal peptide of Hbp mediates targeting to the inner membrane via the SRP pathway. When the SRP pathway is compromised, SecB can prevent, to a certain extent, the mislocalization of pre-pro-Hbp. Subsequent translocation across the inner membrane involves the Sec-translocon. Subsequent translocation across the inner membrane involves the Sec-translocon This is the first demonstration of the use of the co-translational SRP pathway for inner membrane targeting of an extracellular protein

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION