Switching base preferences of mismatch cleavage in endonuclease V: an improved method for scanning point mutations.
Switching base preferences of mismatch cleavage in endonuclease V: an improved method for scanning point mutations.
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DOI：
10.1093/nar/gkl916
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发表时间：
2007
影响因子：
14.9
通讯作者：
Cao W
中科院分区：
文献类型：
--
作者：
Gao H;Huang J;Barany F;Cao W
核酸内切酶V（endo V）识别DNA中广泛的畸变，例如脱氨基碱基或错配。它在第二个磷酸二酯键3′处将DNA切割成脱氨基碱基或错配碱基。从海栖热袍菌获得的内切核酸酶V在某些序列背景下优先切割嘌呤错配。核酸内切酶V已与高保真DNA连接酶结合，以开发用于突变扫描的酶促方法。定点突变体的生化筛选确定了基序III和IV中的突变体，这些突变体改变了错配切割中的碱基偏好。最重要的是，在Y80位置处的单个丙氨酸取代将该酶基本上转变为C特异性错配核酸内切酶，其识别并切割A/C、C/A、T/C、C/T，甚至先前难治的C/C错配。Y80 A还可以检测K-ras癌基因中的G13 D突变，这是一种嵌入在G/C丰富序列背景中的A/C错配，以前使用野生型endo V无法获得。endo V中的蛋白质工程可能转化为突变识别和癌症突变扫描的更好工具。
Endonuclease V (endo V) recognizes a broad range of aberrations in DNA such as deaminated bases or mismatches. It nicks DNA at the second phosphodiester bond 3′ to a deaminated base or a mismatch. Endonuclease V obtained from Thermotoga maritima preferentially cleaves purine mismatches in certain sequence context. Endonuclease V has been combined with a high-fidelity DNA ligase to develop an enzymatic method for mutation scanning. A biochemical screening of site-directed mutants identified mutants in motifs III and IV that altered the base preferences in mismatch cleavage. Most profoundly, a single alanine substitution at Y80 position switched the enzyme to essentially a C-specific mismatch endonuclease, which recognized and cleaved A/C, C/A, T/C, C/T and even the previously refractory C/C mismatches. Y80A can also detect the G13D mutation in K-ras oncogene, an A/C mismatch embedded in a G/C rich sequence context that was previously inaccessible using the wild-type endo V. This investigation offers insights on base recognition and active site organization. Protein engineering in endo V may translate into better tools in mutation recognition and cancer mutation scanning.
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DNA对固体支持的化学切割反应：在突变检测中的应用。
DOI：
10.1186/1472-6769-3-1
发表时间：
2003-05-13
期刊：
BMC chemical biology
影响因子：
--
作者：
Bui, Chinh T.;Lambrinakos, Andreana;Babon, Jeffrey J.;Cotton, Richard G. H.
通讯作者：
Cotton, Richard G. H.
影响因子：
14.9
作者：
Pincas, H;Pingle, MR;Barany, F
通讯作者：
Barany, F
DOI：
10.1073/pnas.0405913101
发表时间：
2004-10-05
影响因子：
11.1
作者：
Rand, TA;Ginalski, K;Wang, XD
通讯作者：
Wang, XD
影响因子：
14.9
作者：
Handa, P;Acharya, N;Varshney, U
通讯作者：
Varshney, U
影响因子：
2.9
作者：
Hecker, KH;Taylor, PD;Gjerde, DT
通讯作者：
Gjerde, DT