Defining the role of the Escherichia coli chaperone SecB using comparative proteomics.
To improve understanding and identify novel substrates of the cytoplasmic chaperone SecB in Escherichia coli, we analyzed a secB null mutant using comparative proteomics. The secB null mutation did not affect cell growth but caused significant differences at the proteome level. In the absence of SecB, dynamic protein aggregates containing predominantly secretory proteins accumulated in the cytoplasm. Unprocessed secretory proteins were detected in radiolabeled whole cell lysates. Furthermore, the assembly of a large fraction of the outer membrane proteome was slowed down, whereas its steady state composition was hardly affected. In response to aggregation and delayed sorting of secretory proteins, cytoplasmic chaperones DnaK, GroEL/ES, ClpB, IbpA/B, and HslU were up-regulated severalfold, most likely to stabilize secretory proteins during their delayed translocation and/or rescue aggregated secretory proteins. The SecB/A dependence of 12 secretory proteins affected by the secB null mutation (DegP, FhuA, FkpA, OmpT, OmpX, OppA, TolB, TolC, YbgF, YcgK, YgiW, and YncE) was confirmed by "classical" pulse-labeling experiments. Our study more than triples the number of known SecB-dependent secretory proteins and shows that the primary role of SecB is to facilitate the targeting of secretory proteins to the Sec-translocase.
Baars
Louise
L
Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University SE-106 91 Stockholm, Sweden.
Ytterberg
A Jimmy
AJ
Drew
David
D
Wagner
Samuel
S
Thilo
Claudia
C
van Wijk
Klaas Jan
KJ
de Gier
Jan-Willem
JW
D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D019295:Computational Biology; D003593:Cytoplasm / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism; D005434:Flow Cytometry; D007091:Image Processing, Computer-Assisted; D015151:Immunoblotting; D047468:Immunoprecipitation; D013058:Mass Spectrometry; D018832:Molecular Chaperones / Q000378:metabolism*; D009154:Mutation; D010641:Phenotype; D011485:Protein Binding; D040901:Proteomics / Q000379:methods*
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; C057842:SecB protein, Bacteria