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Biochemistry. 1998 Aug 11;37(32):11301-8.

Linkage between oligomerization and DNA binding in Drosophila doublesex proteins.

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The Rosenstiel Center,The Graduate Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02254-9110, USA.


The doublesex gene of Drosophila melanogaster encodes DSXM protein in males and DSXF protein in females. Dimers of each protein bind a DNA site from which DSXM represses and DSXF activates transcription. Amino acids 1-397 are identical between the proteins and include a domain (DBD) for both DNA binding and protein oligomerization. The remaining nonhomologous and therefore sex-specific C-termini include an essential part of a second oligomerization domain. We have used mobility shift assays to investigate the effects these three oligomerization domains (DBD and two sex-specific) have on DSX dimerization and DNA binding. The intrinsic DNA binding affinities of DSXM and DSXF dimers are indistinguishable from each other (0.17 +/- 0.04 nM) and slightly lower than that of DBD dimers (0.48 nM). In contrast, the dimerization dissociation constants of DSXM (0.05 +/- 0.02 nM) and DSXF (0.16 +/- 0.05 nM) are slightly different, but 4 orders of magnitude lower than that of DBD (430 nM). Thus sequences outside of DBD, presumably the sex-specific oligomerization domains, have substantial effects on apparent DNA binding affinity through thermodynamically linked effects on dimerization of full-length proteins. Further, when two DNA binding sites are adjacent, DBD dimers show no binding cooperativity, whereas full-length dimers bind with 2-fold different cooperativity (DSXF, k12 = 2.6; DSXM, k12 = 5.4). This suggests that the sex-specific domains may have a second effect on DNA binding, namely, an effect on binding cooperativity that depends on the number and arrangement of DNA sites.

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