Maturation and localization of the TolB protein required for colicin import.
The tolB gene has been shown previously to encode two proteins of 47.5 kDa (TolB) and 43 kDa (TolB*). To explain the presence of these two forms, two hypotheses have been proposed: TolB might be posttranslationally processed to TolB*, or an internal in-frame translation initiation resulting in TolB* may occur (S. K. Levengood and R. E. Webster, J. Bacteriol. 171:6600-6609, 1989). To address this question, TolB was tagged by inserting in its C-terminal region an epitope recognized by monoclonal antibody 1C11 without altering the function of TolB. It was then demonstrated that the functional protein corresponded to TolB*, the mature periplasmic protein, and that TolB was its precursor form, which was observed only when the protein was overexpressed. These two forms were purified by immunoprecipitation, and their N-terminal sequences were determined. An antibody directed against TolB was raised, which confirmed the results obtained with the tagged TolB.
Isnard
M
M
Laboratoire d'Ingénierie et de Dynamique des Systèmes Membranaires, Centre National de la Recherche Scientifique (CNRS), Marseille, France.
Rigal
A
A
Lazzaroni
J C
JC
Lazdunski
C
C
Lloubes
R
R
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D001483:Base Sequence; D001692:Biological Transport; D002451:Cell Compartmentation; D003087:Colicins / Q000378:metabolism; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D033903:Periplasmic Proteins; D011498:Protein Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D017421:Sequence Analysis
D001426:Bacterial Proteins; D003087:Colicins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D033903:Periplasmic Proteins; D011498:Protein Precursors; C062094:tolB protein, E coli