I. EXPRESSION, PURIFICATION, AND COMPARISON OF DE NOVO AND MAINTENANCE METHYLATION*
I. EXPRESSION, PURIFICATION, AND COMPARISON OF DE NOVO AND MAINTENANCE METHYLATION*
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I. 从头甲基化和维持甲基化的表达、纯化和比较*
DOI：
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发表时间：
1999
期刊：
影响因子：
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通讯作者：
R. Roberts
中科院分区：
文献类型：
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作者：
S. Pradhan;A. Bacolla;R. Wells;R. Roberts
（kcat）在poly（dI-dC）zpoly（dI-dC）上为131 - 237 h 21，在未甲基化DNA上为1.2 - 2.3 h 21，在半甲基化DNA上为8.3 - 49 h 21。DNA（Km）和S-腺苷-L-甲硫氨酸（Met）（Km）的米氏常数范围分别为0.33 - 1.32和2.6 - 7.2 mM，而kcat/Km的比值范围为3.9 - 44（poly（dI-dC）zpoly（dI-dC）为237 - 336）3 10 6 M 21 h 21。相对于未甲基化的DNA，该酶对半甲基化的DNA的偏好是7 - 21倍。半甲基化DNA上的k cat值显示出2 - 3倍的差异，这取决于哪条链被预甲基化。此外，人DNMT 1与含有5-氟-CNG的底物形成共价复合物，表明底物特异性延伸超出了典型的CG二核苷酸。这些结果表明，除了维持甲基化，人DNMT 1也可以进行从头和非CG甲基转移酶的活动在体内。
(kcat) was 131‐237 h 21 on poly(dI-dC)zpoly(dI-dC), 1.2‐2.3 h 21 on unmethylated DNA, and 8.3‐ 49 h 21 on hemimethylated DNA. The Michaelis constants for DNA (K m ) and S-adenosyl-L-methionine (AdoMet) (Km ) ranged from 0.33‐1.32 and 2.6 ‐7.2 mM, respectively, whereas the ratio of kcat/Km ranged from 3.9 to 44 (237‐336 for poly(dI-dC)zpoly(dI-dC)) 3 10 6 M 21 h 21 . The preference of the enzyme for hemimethylated, over unmethylated, DNA was 7‐21-fold. The values of k cat on hemimethylated DNAs showed a 2‐3-fold difference, depending upon which strand was pre-methylated. Furthermore, human DNMT1 formed covalent complexes with substrates containing 5-fluoro-CNG, indicating that substrate specificity extended beyond the canonical CG dinucleotide. These results show that, in addition to maintenance methylation, human DNMT1 may also carry out de novo and non-CG methyltransferase activities in vivo.
DOI：
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发表时间：
1998
期刊：
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影响因子：
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作者：
R. Wells;S. Warren;M. Sarmiento
通讯作者：
R. Wells;S. Warren;M. Sarmiento