Export and localization of N-terminally truncated derivatives of Escherichia coli K-12 outer membrane protein PhoE.
To identify export and sorting information in outer membrane protein PhoE of Escherichia coli K-12, a set of deletions was created, resulting in the removal of N-terminal amino acids of the mature protein. Pulse-chase experiments revealed that some mutant proteins were slowly or not at all processed, but there was not correlation between processing rate and the extent of the deletions. The unprocessed precursors were accessible to trypsin in the periplasm showing that processing by leader peptidase rather than translocation is affected by these deletions. The results show that no specific sequences in the N-terminal part of the mature PhoE protein are required for translocation through the inner membrane. The capability of the processed mutant proteins to assemble into the outer membrane was correlated to the exten of the deletions. Thus, mutants which lack up to amino acid residue 14 are normally incorporated into the outer membrane. Larger deletions which removed the first postulated membrane-spanning fragment of the protein affected the efficiency of assembly: in addition to trimers of the protein in the outer membrane, also monomers were detected in the periplasm. If the deletions extended C-terminally to residue 48, only monomeric forms of the proteins were found in the periplasm.
Bosch
D
D
Department of Molecular Cell Biology, State University Utrecht, The Netherlands.
Voorhout
W
W
Tommassen
J
J
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics; D004274:DNA, Recombinant; D004492:Edetic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D006651:Histocytochemistry; D007118:Immunoassay; D007473:Ion Channels; D007700:Kinetics; D046911:Macromolecular Substances; D008565:Membrane Proteins; D009154:Mutation; D010957:Plasmids; D018272:Porins; D011498:Protein Precursors / Q000378:metabolism; D012697:Serine Endopeptidases; D014169:Transformation, Bacterial; D014357:Trypsin / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial; D004274:DNA, Recombinant; D007473:Ion Channels; D046911:Macromolecular Substances; D008565:Membrane Proteins; D018272:Porins; D011498:Protein Precursors; D004492:Edetic Acid; D010450:Endopeptidases; D012697:Serine Endopeptidases; D014357:Trypsin; C026515:type I signal peptidase