Structure of the Escherichia coli Fis-DNA complex probed by protein conjugated with 1,10-phenanthroline copper(I) complex.
The Escherichia coli Fis (factor for inversion stimulation) protein functions in many diverse biological systems including recombination, transcription, and DNA replication. Although Fis is a site-specific DNA-binding protein, it lacks a well-defined consensus recognition sequence. The electrophoretic mobility of Fis-DNA complexes, along with considerations of the Fis crystal structure, indicates that significant deformation of DNA occurs upon Fis binding. To investigate the structure of Fis-DNA complexes, the chemical nuclease 1,10-phenanthroline-copper complex (OP-Cu) has been linked to four specific sites within the Fis DNA-binding domain. Two of these Fis-OP derivatives were active in cleaving DNA. The scission patterns obtained on four different Fis binding sites indicate that Fis positions itself on these highly divergent DNA sequences in a very similar fashion. The patterns of cleavage of a derivative at Asn-98 generally support a model of a Fis-DNA complex that contains specific bends within the core-recognition sequence. Data from a second Fis-OP derivative at Asn-73 provides evidence for greater wrapping of flanking DNA around the sides of the Fis protein than was previously postulated. The cleavage efficiency of flanking segments varies, suggesting that the extent of DNA wrapping is sequence dependent. Specific amino acids on Fis are implicated in promoting this DNA wrapping.
Pan
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Molecular Biology Institute, University of California, Los Angeles 90024-1737.
Feng
J A
JA
Finkel
S E
SE
Landgraf
R
R
Sigman
D
D
Johnson
R C
RC
D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics; D003432:Cross-Linking Reagents; D004269:DNA, Bacterial / Q000737:chemistry* / Q000235:genetics; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D029968:Escherichia coli Proteins; D035021:Factor For Inversion Stimulation Protein; D034982:Integration Host Factors; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D010618:Phenanthrolines; D011487:Protein Conformation
D002352:Carrier Proteins; D003432:Cross-Linking Reagents; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D035021:Factor For Inversion Stimulation Protein; D034982:Integration Host Factors; D010618:Phenanthrolines; C440502:integration host factor, E coli; C014741:bis(1,10-phenanthroline)copper(2+) ion