Kaur K, Rohozinski J, Goorha R
Department of Virology and Molecular Biology, St Jude Children's Research Hospital, Memphis, TN 38105, USA.
J Gen Virol. 1995 Aug;76 ( Pt 8):1937-43. doi: 10.1099/0022-1317-76-8-1937.
Cytosine DNA methyltransferases (MTases) first recognize specific nucleotide sequences and then transfer a methyl group from S-adenosylmethionine to cytosine. This division of function is reflected in five highly conserved motifs shared by cytosine MTases. The region containing the first four motifs is responsible for the catalytic function whereas the region containing the fifth motif V provides specificity of binding to DNA. In at least one case, two separate proteins, one containing the first four motifs and the second containing the last motif combine to provide full functional activity. In the frog virus 3 (FV3) genome we have identified an open reading frame (ORF) whose deduced amino acid (aa) sequence contains motifs characteristic of prokaryotic as well as eukaryotic MTases. The ORF consists of 642 bp which codes for a protein of 214 aa with a predicted molecular mass of 24.8 kDa. This ORF contains the first four highly conserved motifs of cytosine MTases but the fifth motif, responsible for DNA binding specificity, is missing. Presumably, FV3 MTase is composed of two subunits. Northern blot analysis showed that the putative MTase ORF is transcribed into two transcripts belonging to the delayed-early class of FV3 messages. These two transcripts appear to be initiated at two different start sites but terminate in the same 3' region of the gene. The transcription start sites are not preceded by any known promoter sequences, but two regions of hyphenated dyad symmetry are present at the 3' end of the message. A protein with a molecular mass of approximately 28 kDa was synthesized by a rabbit reticulocyte lysate programmed with capped runoff transcripts from the cloned gene, suggesting that the ORF can be transcribed into a message coding for a viral protein. Overall, our results suggest that we have identified a gene for a subunit of MTase in the FV3 genome.
胞嘧啶DNA甲基转移酶（MTases）首先识别特定的核苷酸序列，然后将甲基从S-腺苷甲硫氨酸转移至胞嘧啶。这种功能划分体现在胞嘧啶MTases共有的五个高度保守的基序中。包含前四个基序的区域负责催化功能，而包含第五个基序V的区域则提供与DNA结合的特异性。至少在一种情况下，两种不同的蛋白质，一种包含前四个基序，另一种包含最后一个基序，结合起来提供完整的功能活性。在蛙病毒3（FV3）基因组中，我们鉴定出一个开放阅读框（ORF），其推导的氨基酸（aa）序列包含原核生物和真核生物MTases的特征基序。该ORF由642 bp组成，编码一个214 aa的蛋白质，预测分子量为24.8 kDa。这个ORF包含胞嘧啶MTases的前四个高度保守基序，但缺少负责DNA结合特异性的第五个基序。推测FV3 MTase由两个亚基组成。Northern印迹分析表明，推定的MTase ORF转录成两种属于FV3延迟早期类信息的转录本。这两种转录本似乎在两个不同的起始位点起始，但在基因的同一3'区域终止。转录起始位点之前没有任何已知的启动子序列，但在信息的3'端存在两个带连字符的二元对称区域。用来自克隆基因的加帽径流转录本编程的兔网织红细胞裂解物合成了一种分子量约为28 kDa的蛋白质，这表明该ORF可以转录成编码病毒蛋白的信息。总体而言，我们的结果表明我们在FV3基因组中鉴定出了一个MTase亚基的基因。
