Single strand DNA specificity analysis of human nucleoside diphosphate kinase B

Nucleoside diphosphate kinases (NDP kinases) form a family of oligomeric en zymes present in all organisms, Eukaryotic NDP kinases are hexamers compose d of identical subunits (approximate to 17 kDa), A distinctive property of human NDPK-B encoded by the gene nm23-H-2 is its ability to stimulate the g ene transcription. This property is independent of its catalytic activity a nd is possibly related to the role of this protein in cellular events inclu ding differentiation and tumor metastasis. In this paper, we report the fir st characterization of human NDPK-B.DNA complex formation using a filter-bi nding assay and fluorescence spectroscopy. We analyzed the binding of sever al oligonucleotides mimicking the promoter region of the c-myc oncogene inc luding variants in sequence, structure, and length of both strands, We show that NDPK-B binds to single-stranded oligonucleotides in a nonsequence spe cific manner, but that it exhibits a poor binding activity to double-strand ed oligonucleotides. This indicates that the specificity of recognition to DNA is a function of the structural conformation of DNA rather than of its specific sequence. Moreover, competition experiments performed with all nuc leotides provide evidence for the contribution of the six active sites in t he DNA-protein complex formation. We propose a mechanism through which huma n NDPK-B could stimulate transcription of c-myc or possibly other genes inv olved in cellular differentiation.