The Campylobacter jejuni PEB1a adhesin is an aspartate/glutamate-binding protein of an ABC transporter essential for microaerobic growth on dicarboxylic amino acids.
The PEB1a protein of the gastrointestinal pathogen Campylobacter jejuni mediates interactions with epithelial cells and is an important factor in host colonization. Cell fractionation and immunoblotting showed that PEB1a is most abundant in the periplasm of C. jejuni, and is detectable in the culture supernatant but not in the inner or outer membrane. The protein is homologous with periplasmic-binding proteins associated with ABC transporters and we show by fluorescence spectroscopy that purified recombinant PEB1a binds L-aspartate and L-glutamate with sub microM K(d) values. Binding of L-14C-aspartate or L-14C-glutamate was strongly out-competed by excess unlabelled aspartate or glutamate but only poorly by asparagine and glutamine. A mutant in the Cj0921c gene, encoding PEB1a, was completely unable to transport 5 microM L-14C-glutamate and showed a large reduction (approximately 20-fold) in the rate of L-14C-aspartate transport compared with the wild type. Although microaerobic growth of this mutant was little affected in complex media, growth on aspartate or glutamate in defined media was completely prevented, whereas growth with serine was similar to wild type. 1H-NMR analysis of the culture supernatants of the Cj0921c mutant showed some utilization of aspartate but not glutamate, consistent with the transport data. It is concluded that in addition to the established role of PEB1a as an adhesin, the PEB1 transport system plays a key role in the utilization of aspartate and glutamate, which may be important in vivo carbon sources for this pathogen.
Leon-Kempis
Maria del Rocio
Mdel R
Department of Molecular Biology and Biotechnology, University of Sheffield, Firth Court, Western Bank, Sheffield S10 2TN, UK.
Guccione
Edward
E
Mulholland
Francis
F
Williamson
Michael P
MP
Kelly
David J
DJ
D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D000332:Aerobiosis / Q000502:physiology*; D000600:Amino Acids, Dicarboxylic / Q000378:metabolism*; D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics / Q000378:metabolism*; D001224:Aspartic Acid / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D016123:Campylobacter jejuni / Q000166:cytology / Q000235:genetics / Q000502:physiology*; D003470:Culture Media / Q000737:chemistry; D018698:Glutamic Acid / Q000378:metabolism*; D006801:Humans; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013347:Subcellular Fractions / Q000737:chemistry
D018528:ATP-Binding Cassette Transporters; D000600:Amino Acids, Dicarboxylic; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D003470:Culture Media; C489977:PEB1 protein, Campylobacter; D011994:Recombinant Proteins; D001224:Aspartic Acid; D018698:Glutamic Acid