Neisseria gonorrhoeae FitA Interacts with FitB To Bind DNA through Its Ribbon−Helix−Helix Motif

• Published in: Biochemistry (Easton) 2005-09, Vol.44 (37), p.12515-12524
• Main Authors: Wilbur, J. Scott, Chivers, Peter T, Mattison, Kirsten, Potter, Laura, Brennan, Richard G, So, Magdalene
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Bacterial Proteins - chemistry; Bacterial Proteins - metabolism; Base Sequence; Carrier Proteins - chemistry; Carrier Proteins - metabolism; Consensus Sequence; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - metabolism; Molecular Sequence Data; Neisseria gonorrhoeae; Neisseria gonorrhoeae - metabolism; Oligonucleotide Probes; Polymerase Chain Reaction; Protein Conformation; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - metabolism; Sequence Alignment; Sequence Homology, Amino Acid
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi0511080

The fit locus, encoding two proteins, FitA and FitB, was identified in a genetic screen for Neisseria gonorrhoeae determinants that affect trafficking across polarized epithelial cells. To better understand how the locus may control these activities, we have undertaken a biochemical analysis of FitA and FitB. FitA is a DNA-binding protein with a putative ribbon−helix−helix (RHH) motif. Purified FitA forms a homodimer that binds a 150 bp fit promoter sequence containing the translational start site. A putative β strand mutant of FitA, FitA(R7A), is unable to bind this DNA, supporting further that FitA is a RHH protein. FitB interacts with FitA to form a 98 kDa complex. FitA/B binds DNA with a 38-fold higher affinity than the FitA homodimer. In DNase I footprint assays, FitA/B protects a 62-bp region within the fit promoter containing the predicted −10 sequence and an 8-bp inverted repeat, TGCTATCA-N12-TGATAGCA. FitA/BHis is able to bind to either half-site alone with high affinity.