DNA-dependent and DNA-independent associations of DNA-binding proteins are important in transcriptional regulation. The analysis of DNA-independent associations frequently relies on assaying protein interaction in the absence of target DNA sequences. We have found that contaminating DNA in protein preparations can stabilize DNA-dependent associations that may appear DNA-independent. Three cellular proteins of 70, 85, and 110 kDa coimmunoprecipitated with the octamer motif-binding protein Oct-2 because of the presence of contaminating DNA in the cell extracts. In addition, heterodimer formation between Oct-1 (or Oct-2) and Pit-1 during protein-affinity chromatography was stabilized by the contaminating DNA. In both instances, these DNA-dependent protein associations were selectively inhibited by ethidium bromide in the precipitation reaction without any evident effect on DNA-independent protein associations. Thus, ethidium bromide may serve as a simple and general indicator of DNA-dependent and DNA-independent protein associations.
[1]
B R Franza,et al.
Association of adenovirus early-region 1A proteins with cellular polypeptides
,
1986,
Molecular and cellular biology.
[2]
E. Harlow,et al.
Antibodies: A Laboratory Manual
,
1988
.
[3]
E. Harlow,et al.
Antibodies specific for the human retinoblastoma protein identify a family of related polypeptides
,
1991,
Molecular and cellular biology.
[4]
C. Verrijzer,et al.
The Oct-1 POU domain mediates interactions between Oct-1 and other POU proteins
,
1992,
Molecular and cellular biology.
[5]
P. Sharp,et al.
A lymphoid-specific protein binding to the octamer motif of immunoglobulin genes
,
1986,
Nature.