The HimA and HimD subunits of integration host factor can specifically bind to DNA as homodimers.
Integration host factor (IHF) is a heterodimeric protein from Escherichia coli which specifically binds to an asymmetric consensus sequence. We have isolated the individual subunits of IHF, HimA and HimD, and show that an active IHF protein can be reconstituted from these subunits. The HimA and HimD polypeptides alone are capable of specifically recognizing the same ihf sequence. The mobilities of the protein-DNA complexes in a gel-retardation assay suggest that the proteins bind as homodimers. The stability of the HimD-DNA complex is approximately 100-fold lower than that of the IHF-DNA complex. The HimA-DNA complex is even less stable and is only observed when a large excess of HimA is used. This instability is possibly due to the inability of HimA to form stable homodimers. By domain swapping between HimA and HimD, we have constructed an IHF fusion protein which has the putative DNA-binding domains of only HimA. This fusion protein forms stable dimers and makes specific protein-DNA complexes with a high efficiency. A comparable fusion protein with only the DNA-binding domains of HimD forms less stable complexes, suggesting that sequence-specific contacts between IHF and the ihf consensus are mainly provided by the HimA subunit.
Zulianello
L
L
Laboratory of Molecular Genetics, Leiden Institute of Chemistry, Leiden University, The Netherlands.
de la Gorgue de Rosny
E
E
van Ulsen
P
P
van de Putte
P
P
Goosen
N
N
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism*; D034982:Integration Host Factors; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D034982:Integration Host Factors; D011993:Recombinant Fusion Proteins