Z-DNA-binding proteins. Identification critically depends on the proper choice of ligands.
We have previously isolated from bull testis three proteins of molecular mass 31, 33, and 58 kDa that we have tentatively characterized as high affinity Z-DNA-binding proteins. This inference was based on their preferential binding to brominated poly(dG-dC).poly(dG-dC) in Z-form as opposed to the unbrominated polynucleotide in B-form (Gut, S. H., Bischoff, M., Hobi, R., and Kuenzle, C. C. (1987) Nucleic Acids Res. 15, 9691-9705). By partial amino acid sequencing we have provisionally identified the 31- and 33-kDa proteins as members of the high mobility group 2 and 1 protein families, respectively, whereas the 58-kDa protein has so far remained unidentified (Christen, Th., Bischoff, M., Hobi, R., and Kuenzle, C. C. (1990) FEBS Lett. 267, 139-141). In the present study, we have critically reassessed the binding specificity of these three proteins by using more natural Z- and B-DNA ligands. As such we chose supercoiled and relaxed DNA minicircles containing a d(CG)7 insert in the Z- and B-conformation, respectively. Filter binding tests and gel retardation assays performed with these ligands showed that the three testis proteins either do not discriminate between Z- and B-DNA (31- and 33-kDa proteins) or even have a preference for B-DNA (58-kDa protein). Therefore, we question the validity of using brominated poly(dG-dC).poly(dG-dC) as an indicator of Z-DNA binding.
Rohner
K J
KJ
Institut für Pharmakologie und Biochemie, Universität Zürich-Irchel, Switzerland.
Hobi
R
R
Kuenzle
C C
CC
D000818:Animals; D000906:Antibodies; D001483:Base Sequence; D002417:Cattle; D004247:DNA / Q000378:metabolism*; D004270:DNA, Circular / Q000378:metabolism; D018105:DNA, Kinetoplast; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009690:Nucleic Acid Conformation; D011089:Polydeoxyribonucleotides / Q000378:metabolism; D013737:Testis / Q000378:metabolism*
D000906:Antibodies; D004270:DNA, Circular; D018105:DNA, Kinetoplast; D004268:DNA-Binding Proteins; D011089:Polydeoxyribonucleotides; C019530:poly(dC-dG); D004247:DNA