Specificity of mammalian Y-box binding protein p50 in interaction with ss and ds DNA analyzed with generic oligonucleotide microchip
Specificity of mammalian Y-box binding protein p50 in interaction with ss and ds DNA analyzed with generic oligonucleotide microchip
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DOI：
10.1016/s0022-2836(02)00937-3
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发表时间：
2002-11-15
影响因子：
5.6
通讯作者：
Mirzabekov, AD
中科院分区：
文献类型：
--
作者：
Zasedateleva, OA;Krylov, AS;Mirzabekov, AD
p50 protein is a member of the Y-box binding transcription factor family and is a counterpart of YB-1 protein. The generic microchip was used to analyze the sequence specificity of p50 binding to single (ss) and double-stranded (ds) oligodeoxyribonucleotides. The generic microchip contained 4096 single-stranded octacleoxyribonucleotides in which all possible core 6-mers (4(6) = 4096) were flanked at their 3' and 5'-ends with degenerated nucleotides. The oligonucleotides were chemically immobilized within polyacrylamide gel pads fixed on a glass slide. The binding of p50 to the generic microchip was shown to be the most specific to ss GGGG motif and then to ss CACC and CATC motifs. GC-rich ds oligonucleotides of the generic microchip, and particularly those containing GGTG/CACC, GATG/CATC, and GTGG/CCAC heterogeneous motifs, were most efficiently destabilized due to interaction with p50. Gel-shift electrophoresis has shown that the protein exhibits much higher binding specificity to 24-mer oligoA-TGGGGG-oligoA containing G-rich 6-mer, in comparison with 24-mer oligoA-AAATAT-oligoA carrying A,T-rich 6-mer in full correspondence with the data obtained with the microchip. Studies of DNA-binding proteins using gel-immobilized ss and ds DNA fragments provide a unique possibility to detect low-affinity complexes of these proteins with short sequence motifs and assess the role of these motifs in sequence-specific interactions with long recognition sites. (C) 2002 Elsevier Science Ltd. All rights reserved.
