DNA-CYTOS-METHYLTRANSFERASE (S)

本申请的长期目标是获得详细的 了解药物的结构、作用机制和调节 人DNA甲基转移酶（E.C.2.1.1.37）。 现有证据有力地证明 这表明，这种催化的DNA合成后修饰 酶在细胞分化和体细胞遗传中起作用。 DNA 甲基化还可能参与异位蛋白质的产生、肿瘤的发生、肿瘤的形成和转移。 起始和肿瘤进展。 我们建议制备和研究毫克定量的均匀 从人胎盘提取的酶 旨在研究这种酶的实验 结构包括确定亚基化学计量和分子 重量和氨基酸序列的部分测定。 纯化的 酶将用于构建和优化体外DNA甲基化 系统 该系统和纯化酶的抗血清将用于 实验以确定可能促进或抑制DNA甲基化的因素 在细胞增殖过程中，以及可能导致 在某些转化细胞系中观察到的DNA低甲基化。 氨基酸序列信息将用于合成 用作杂交探针以帮助分离 甲基转移酶的cDNA克隆。 由此分离的cDNA克隆 将用于确定基因组组织和转录 甲基转移酶基因的调节。

The long-term objectives of this application are to obtain a detailed understanding of the structure, mechanisms of action, and regulation of the human DNA methyltransferase(s) (E.C.2.1.1.37). Available evidence strongly suggests that the postsynthetic modification of DNA catalyzed by this enzyme has a role in cellular differentiation and somatic inheritance. DNA methylation may also be involved in ectopic protein production, tumor initiation, and tumor progression. We propose to prepare and study milligram quantitites of the homogeneous enzyme from human placenta. Experiments designed to study the enzyme structure include the determination of sub-unit stoichiometry and molecular weight, and a partial determination of amino acid sequence. The purified enzyme will be used to construct and optimize an in vitro DNA methylation system. This system and antisera to the purified enzyme will be used in experiments to identify factors that may promote or inhibit DNA methylation during cell proliferation, and factors that may be responsible for the hypomethylation of DNA observed in certain transformed cell lines. The amino acid sequence information will be used to synthesize oligonucleotides for use as hybridization probes to aid in the isolation of a cDNA clone for the methyltransferase. The cDNA clone(s) thus isolated will be used to determine the genomic organization and the transcriptional regulation of the methyltransferase gene(s).