Membrane insertion characteristics of the various transmembrane domains of the Escherichia coli TolQ protein.
The Escherichia coli TolQ protein is a 230-amino acid integral cytoplasmic membrane protein required for the import of group A colicins, for infection by the filamentous phage, and for maintenance of the integrity of the bacterial envelope. TolQ is a polytopic protein with three membrane-spanning regions. The first membrane-spanning region has a 19-residue periplasmic NH2-terminal tail, while the second and third membrane-spanning segments are separated by a short 17-amino acid periplasmic loop. To study the membrane assembly of TolQ, fusions of different membrane-spanning regions were examined for their ability to insert in the absence of functional SecA or the membrane potential. Fusions containing the first membrane-spanning region plus the adjacent cytoplasmic domain and a construct containing the "hairpin loop," formed by the second and third membrane-spanning regions, insert in the absence of functional SecA. The fusion containing the second and third membrane-spanning regions required the membrane potential for insertion while the first membrane-spanning region was able to insert even in the absence of a membrane potential. Taken together, these results suggest that insertion of intact TolQ is not dependent on the Sec system, but does require the membrane potential.
Lewin
T M
TM
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
Webster
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RE
D000251:Adenosine Triphosphatases / Q000378:metabolism; D001386:Azides / Q000494:pharmacology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D015870:Gene Expression / Q000187:drug effects; D008564:Membrane Potentials; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D026901:Membrane Transport Proteins; D008961:Models, Structural; D010446:Peptide Fragments / Q000302:isolation & purification; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D014357:Trypsin
D001386:Azides; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D000069816:SEC Translocation Channels; C060792:tolQ protein, E coli; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D014357:Trypsin; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins