title: Small-angle X-ray scattering and in silico modeling approaches for the accurate functional annotation of an LysR-type transcriptional regulator.
abstract: Xylella fastidiosa is a xylem-limited, Gram-negative phytopathogen responsible for economically relevant crop diseases. Its genome was thus sequenced in an effort to characterize and understand its metabolism and pathogenic mechanisms. However, the assignment of the proper functions to the identified open reading frames (ORFs) of this pathogen was impaired due to a lack of sequence similarity in the databases. In the present work, we used small-angle X-ray scattering and in silico modeling approaches to characterize and assign a function to a predicted LysR-type transcriptional regulator in the X. fastidiosa (XfLysRL) genome. XfLysRL was predicted to be a homologue of BenM, which is a transcriptional regulator involved in the degradation pathway of aromatic compounds. Further functional assays confirmed the structural prediction because we observed that XfLysRL interacts with benzoate and cis,cis-muconic acid (also known as 2E,4E-hexa-2,4-dienedioic acid; hereafter named muconate), both of which are co-factors of BenM. In addition, we showed that the XfLysRL protein is differentially expressed during the different stages of X. fastidiosa biofilm formation and planktonic cell growth, which indicates that its expression responds to a cellular signal that is likely related to the aromatic compound degradation pathway. The assignment of the proper function to a protein is a key step toward understanding the cellular metabolic pathways and pathogenic mechanisms. In the context of X. fastidiosa, the characterization of the predicted ORFs may lead to a better understanding of the cellular pathways that are linked to its bacterial pathogenicity.
authors.affiliation: Centro de Biologia Molecular e Engenharia Genética, Universidade Estadual de Campinas, Campinas, SP, Brazil.
authors.forename: M A S
authors.initials: MA
authors.lastname: Toledo
authors.forename: C A
authors.initials: CA
authors.lastname: Santos
authors.forename: J S
authors.initials: JS
authors.lastname: Mendes
authors.forename: A C
authors.initials: AC
authors.lastname: Pelloso
authors.forename: L L
authors.initials: LL
authors.lastname: Beloti
authors.forename: A
authors.initials: A
authors.lastname: Crucello
authors.forename: M T P
authors.initials: MT
authors.lastname: Favaro
authors.forename: A S
authors.initials: AS
authors.lastname: Santiago
authors.forename: D R S
authors.initials: DR
authors.lastname: Schneider
authors.forename: A M
authors.initials: AM
authors.lastname: Saraiva
authors.forename: D R
authors.initials: DR
authors.lastname: Stach-Machado
authors.forename: A A
authors.initials: AA
authors.lastname: Souza
authors.forename: D B B
authors.initials: DB
authors.lastname: Trivella
authors.forename: R
authors.initials: R
authors.lastname: Aparicio
authors.forename: L
authors.initials: L
authors.lastname: Tasic
authors.forename: A R
authors.initials: AR
authors.lastname: Azzoni
authors.forename: A P
authors.initials: AP
authors.lastname: Souza
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001565:Benzoates / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D018441:Biofilms / Q000187:drug effects / Q000254:growth & development; D003198:Computer Simulation; D004591:Electrophoresis, Polyacrylamide Gel; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D053838:Scattering, Small Angle; D017386:Sequence Homology, Amino Acid; D013011:Sorbic Acid / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D014961:X-Ray Diffraction / Q000379:methods*; D044167:Xylella / Q000235:genetics / Q000378:metabolism / Q000502:physiology
chemical_list: D001426:Bacterial Proteins; D001565:Benzoates; D011994:Recombinant Proteins; C005939:muconic acid; D013011:Sorbic Acid