DNA-bending properties of TF1

¹Department of Biology, University of California, San Diego, La Jolla 92093-0634.

|October 5, 1991

Summary

Bacillus subtilis phage SPO1 transcription factor 1 (TF1) sharply bends DNA, similar to E. coli integration host factor (IHF). TF1

Area of Science:

Background:

* Transcription factor 1 (TF1) from Bacillus subtilis phage SPO1 is a DNA-binding protein.
* TF1 belongs to a family including E. coli HU and integration host factor (IHF).
* TF1's DNA-binding affinity and site selectivity are influenced by substitutions at amino acid 61.

Purpose of the Study:

* To investigate the DNA-bending properties of TF1.
* To determine how amino acid substitutions affect TF1's DNA binding and bending.
* To analyze TF1's interaction with thymine-containing DNA and map binding sites.

Main Methods:

* Gel electrophoretic retardation assay to study DNA binding and bending.
* Hydroxyl radical (.OH) and methylation interference footprinting to map TF1 binding sites.
* Analysis of DNA bending by non-specifically bound TF1.

Main Results:

* TF1 dimer binding induces sharp DNA bending, comparable to E. coli IHF.
* Amino acid substitutions at position 61 affect both DNA binding and bending.
* TF1 binding affinity and selectivity are significantly reduced with thymine (T) instead of (5-hydroxymethyl)uracil (hmUra) in DNA.
* TF1 protects specific DNA segments from hydroxyl radical attack, indicating multiple protein contacts.
* Methylation interference reveals TF1 binding is diminished, but not abolished, by modifications in the central DNA region.

Conclusions:

* TF1 binding induces significant deformation of the DNA B-helix.
* DNA sequence, particularly the presence of hmUra versus T, influences TF1 binding and DNA bending.
* Multiple protein-DNA contacts facilitate DNA wrapping around the TF1 dimer.

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