Neisseria gonorrhoeae FitA interacts with FitB to bind DNA through its ribbon-helix-helix motif.
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 beta 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-N(12)-TGATAGCA. FitA/B(His) is able to bind to either half-site alone with high affinity.
Wilbur
J Scott
JS
Department of Molecular Microbiology and Immunology, School of Medicine, Oregon Health and Sciences University, 3181 SE Sam Jackson Park Road, Portland, Oregon 97239, USA. wilburj@ohsu.edu
Chivers
Peter T
PT
Mattison
Kirsten
K
Potter
Laura
L
Brennan
Richard G
RG
So
Magdalene
M
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D001483:Base Sequence; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D009344:Neisseria gonorrhoeae / Q000378:metabolism*; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D001426:Bacterial Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C505223:FitA protein, Neisseria gonorrhoeae; C505224:FitB protein, Neisseria gonorrhoeae; D015345:Oligonucleotide Probes; D011993:Recombinant Fusion Proteins