Boĭko V P, Svetlova M P, Sazeeva N R, Shestopalov B N, Ariga H, Iguchi-Ariga S M, Tomilin N V
Mol Gen Mikrobiol Virusol. 1997(4):29-31.
Two cDNA clones are sequenced which were isolated from human lymphocyte expression library using Southwestern (DNA-binding) screening with 32P-labeled Alu DNA in the presence of 100-fold excess of unbalanced poly (dI-dC). In one of the sequenced clones (vb22) an open reading frame (ORF) is detected, encoding protein with a new potential DNA-binding (zink-finger) domain, and DNA-binding activity of the protein is directly confirmed after its expression (as GST-fusion protein) in Escherichia coli. The other sequenced clone (wa12) is partially homologous to 15EST sequences present in GENBANK (April, 1996) and cloned from very different human tissues. Connection of these 15 overlapping GENBANK sequences resulted in a longer sequence covering wa12 and having ORF potentially encoding a new 10 kDa polypeptide without any apparent DNA-binding domains. This connected sequence as well as wa12 sequence having only 65 amino acids ORF are unrecognizable by computer software as the protein-coding regions, and we suppose that wa12 transcripts possess DNA-binding activity. Homopyrimidine blocks in RNA longer than 12 nucleotides are known to bind mirror duplex DNA sequences to form triplexes whose stability is comparable to that of protein-DNA complexes, and human promoters contain many such blocks.
对两个cDNA克隆进行了测序，它们是从人淋巴细胞表达文库中分离出来的，采用的是在存在100倍过量不平衡聚（dI-dC）的情况下，用32P标记的Alu DNA进行Southwestern（DNA结合）筛选。在其中一个测序克隆（vb22）中检测到一个开放阅读框（ORF），其编码的蛋白质具有一个新的潜在DNA结合（锌指）结构域，并且该蛋白质在大肠杆菌中表达（作为GST融合蛋白）后，其DNA结合活性得到了直接证实。另一个测序克隆（wa12）与GENBANK（1996年4月）中存在的15个EST序列部分同源，并且是从非常不同的人类组织中克隆出来的。将这15个重叠的GENBANK序列连接起来，得到了一个更长的序列，该序列覆盖了wa12，并且具有潜在编码一个新的10 kDa多肽的ORF，而没有任何明显的DNA结合结构域。这个连接后的序列以及只有65个氨基酸ORF的wa12序列，计算机软件无法将其识别为蛋白质编码区，并且我们推测wa12转录本具有DNA结合活性。已知长度超过12个核苷酸的RNA中的同嘧啶区段会与镜像双链DNA序列结合形成三链体，其稳定性与蛋白质-DNA复合物相当，并且人类启动子包含许多这样的区段。
