The Salmonella enterica serovar Typhi LeuO global regulator forms tetramers: residues involved in oligomerization, DNA binding, and transcriptional regulation

• Published in: Journal of bacteriology 2014-06, Vol.196 (12), p.2143-2154
• Main Authors: Guadarrama, Carmen, Medrano-López, Abraham, Oropeza, Ricardo, Hernández-Lucas, Ismael, Calva, Edmundo
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacteriology; Binding sites; Biochemistry; Deoxyribonucleic acid; DNA; DNA, Bacterial - metabolism; E coli; Escherichia coli; Gene Deletion; Gene expression; Gene Expression Regulation, Bacterial - physiology; Gram-negative bacteria; Molecular Sequence Data; Mutagenesis, Site-Directed; Protein Binding; Protein Conformation; Salmonella; Salmonella enterica; Salmonella typhi - genetics; Salmonella typhi - metabolism; Transcription, Genetic
• ISSN: 0021-9193; 1098-5530
• DOI: 10.1128/JB.01484-14

LeuO is a LysR-type transcriptional regulator (LTTR) that has been described to be a global regulator in Escherichia coli and Salmonella enterica, since it positively and negatively regulates the expression of genes involved in multiple biological processes. LeuO is comprised of an N-terminal DNA-binding domain (DBD) with a winged helix-turn-helix (wHTH) motif and of a long linker helix (LH) involved in dimerization that connects the DBD with the C-terminal effector-binding domain (EBD) or regulatory domain (RD; which comprises subdomains RD-I and RD-II). Here we show that the oligomeric structure of LeuO is a tetramer that binds with high affinity to DNA. A collection of single amino acid substitutions in the LeuO DBD indicated that this region is involved in oligomerization, in positive and negative regulation, as well as in DNA binding. Mutants with point mutations in the central and C-terminal regions of RD-I were affected in transcriptional activation. Deletion of the RD-II and RD-I C-terminal subdomains affected not only oligomerization but also DNA interaction, showing that they are involved in positive and negative regulation. Together, these data demonstrate that not only the C terminus but also the DBD of LeuO is involved in oligomer formation; therefore, each LeuO domain appears to act synergistically to maintain its regulatory functions in Salmonella enterica serovar Typhi.