Escherichia coli periplasmic chaperone FaeE is a homodimer and the chaperone-K88 subunit complex is a heterotrimer.
The interaction of FaeE, a periplasmic chaperone involved in K88 biosynthesis, and the major fimbrial subunit FaeG was investigated. The genes encoding the two proteins were subcloned together in the expression vector pINIIIA1. Cells expressing the subcloned genes accumulated in their periplasm a complex of FaeE and FaeG. This complex was purified by isoelectric focusing and anion-exchange fast-protein liquid chromatography. SDS-PAGE, native gel electrophoresis, immunoblotting and determination of the N-terminal amino acid sequences and the molar ratio of the N-terminal amino acid residues revealed that the complex is a heterotrimer consisting of two molecules of FaeE and one molecule of FaeG. The periplasmic chaperone FaeE was purified from the periplasm of cells expressing only the subcloned faeE gene. Gel filtration, protein cross-linking analysis and a biophysical approach in which the rotation diffusion coefficient of the purified FaeE was determined led to the conclusion that the native FaeE chaperone is a homodimer.
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Department of Molecular Microbiology, Faculty of Biology, Vrije Universiteit, Amsterdam, The Netherlands.
Visschers
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RW
de Graff
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FK
Oudega
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D000942:Antigens, Bacterial; D000954:Antigens, Surface / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins; D036781:Fimbriae Proteins; D005798:Genes, Bacterial; D007525:Isoelectric Focusing; D046911:Macromolecular Substances; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D005810:Multigene Family; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism
D000942:Antigens, Bacterial; D000954:Antigens, Surface; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C026888:K88 antigen, E coli; D046911:Macromolecular Substances; D018832:Molecular Chaperones; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; C070012:faeE protein, E coli; D036781:Fimbriae Proteins