Characterization, localization and transmembrane organization of the three proteins PrtD, PrtE and PrtF necessary for protease secretion by the gram-negative bacterium Erwinia chrysanthemi.
Erwinia chrysanthemi, a Gram-negative phythopathogenic bacterium, secretes two related extracellular metalloproteases, B and C, which do not have N-terminal signal sequences. The specific pathway by which they are secreted, which has been reconstituted in Escherichia coli, comprises three proteins -- PrtD, PrtE and PrtF. Hybrid proteins containing segments of these proteins fused to the C-terminus of protease B were purified and used to immunize rabbits. The antisera thus obtained were used to study the location and membrane topology of the three proteins. PrtD and PrtE were found to cofractionate almost exclusively with the cytoplasmic membrane, whereas PrtF was found to co-fractionate mostly with the outer membrane. Proteinase K accessibility experiments as well as sequence data lead us to propose that PrtF has one or both ends exposed to the periplasm, that PrtE has one transmembrane segment with its amino-terminus facing the cytoplasm and its C-terminal hydrophilic domain exposed to the periplasm, and that PrtD has six transmembrane segments with its N-terminus and its C-terminal hydrophilic domain in the cytoplasm.
Delepelaire
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P
Unité de Génétique Moléculaire, URA CNRS 1149, Institut Pasteur, Paris, France.
Wandersman
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C
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D003001:Cloning, Molecular; D016974:Dickeya chrysanthemi / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008666:Metalloendopeptidases / Q000378:metabolism*; D008970:Molecular Weight; D010957:Plasmids; D013104:Spheroplasts / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D008565:Membrane Proteins; D008666:Metalloendopeptidases