ABSTRACT
The oligopeptide-binding protein, OppA, binds and ushers oligopeptide substrates to the membrane-associated oligopeptide permease (Opp), a multi-component ABC-type transporter involved in the uptake of oligopeptides expressed by several bacterial species. In the present study, we report the cloning, purification, refolding and conformational analysis of a recombinant OppA protein derived from Xanthomonas axonopodis pv. citri (X. citri), the etiological agent of citrus canker. The oppA gene was expressed in Escherichia coli BL21 (DE3) strain under optimized inducing conditions and the recombinant protein remained largely insoluble. Solubilization was achieved following refolding of the denatured protein. Circular dichroism analysis indicated that the recombinant OppA protein preserved conformational features of orthologs expressed by other bacterial species. The refolded recombinant OppA represents a useful tool for structural and functional analyses of the X. citri protein.
Subject(s)
Protein Folding , Bacterial Proteins/isolation & purification , Membrane Transport Proteins/isolation & purification , Carrier Proteins/metabolism , Xanthomonas axonopodis/genetics , Amino Acid Sequence , Base Sequence , Computational Biology/methods , Circular Dichroism , Cloning, Molecular , Escherichia coli/genetics , Molecular Sequence Data , Operon , Plasmids , Protein Conformation , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Bacterial Proteins/chemistry , Carrier Proteins/genetics , Carrier Proteins/isolation & purification , Xanthomonas axonopodis/metabolismABSTRACT
The oligopeptide-binding protein, OppA, ushers oligopeptide substrates to the membrane-associated oligopeptide permease (Opp), a multi-component ABC-type transporter involved in the uptake of oligopeptides by several bacterial species. In the present study, we report a structural model and an oligopeptide docking analysis of the OppA protein expressed by Xanthomonas axonopodis pv. citri (X. citri), the etiological agent of citrus canker. The X. citri OppA structural model showed a conserved three-dimensional structure, irrespective of the low amino acid identities with previously defined structures of Bacillus subtilis and Salmonella typhimurium orthologs. Oligopeptide docking analysis carried out with the proposed model indicated that the X. citri OppA preferentially binds tri- and tetrapeptides. The present study represents the first structural analysis of an OppA ortholog expressed by a phytopathogen and contributes to the understanding of the physiology and nutritional strategies of X. citri.
Subject(s)
Lipoproteins/chemistry , Oligopeptides/metabolism , Bacterial Proteins/chemistry , Carrier Proteins/chemistry , Xanthomonas/metabolism , Amino Acid Sequence , Binding Sites , Plant Diseases/microbiology , Ligands , Models, Molecular , Molecular Sequence Data , Protein ConformationABSTRACT
Mandibulofacial dysostosis is readily recognized on the basis of a characteristic facial appearance caused by hard and soft tissue abnormalities of the face, including malformations of the ear. Generally, the abnormality is symmetrical. The psychological and social stigma associated with severe facial deformity makes this syndrome one of the most challenging reconstructive problems presented to the craniomaxillofacial surgeon.