Identification and characterization of two forms of mouse MUTYH proteins encoded by alternatively spliced transcripts
Identification and characterization of two forms of mouse MUTYH proteins encoded by alternatively spliced transcripts
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DOI：
10.1093/nar/gkh214
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发表时间：
2004-01-01
影响因子：
14.9
通讯作者：
Nakabeppu, Y
中科院分区：
文献类型：
--
作者：
Ichinoe, A;Behmanesh, M;Nakabeppu, Y
通过选择性剪接产生的小鼠Mutyh mRNA有三种类型（a、B和c型），并且B型mRNA在所检查的大多数组织中是这三种类型中的主要形式。c型mRNA在脑组织中的表达水平较高。预期a型和B型mRNA编码57.7 kDa蛋白（MUTYH α），而c型mRNA具有部分不同的开放阅读框，编码50.2 kDa蛋白（MUTYH β）。B和c型mRNA的体外翻译分别产生50 kDa MUTYH α和47 kDa MUTYH β。在野生型胚胎干（ES）细胞或从野生型小鼠制备的胸腺细胞中检测到MUTYH α和MUTYH β，但在MUTYH缺失的ES细胞和从MUTYH缺失的小鼠制备的胸腺细胞中均未检测到MUTYH α和MUTYH β。在野生型ES细胞中，MUTYH α和MUTYH β主要定位于细胞核中，一些定位于线粒体中。重组MUTYH α和β在大肠杆菌中表达为与硫氧还蛋白的融合蛋白，但只有MUTYH α是部分可溶的，因此可以纯化。重组MUTYH α具有DNA糖基化酶活性，可切割腺嘌呤对8-氧代鸟嘌呤和鸟嘌呤，但没有AP裂解酶活性。
There are three types of mouse Mutyh mRNAs (type a, b and c) generated by alternative splicing, and type b mRNA is a major form among the three in most of the tissues examined. The level of type c mRNA is relatively high in brain. Type a and b mRNAs were expected to encode 57.7 kDa protein (MUTYHalpha), while type c mRNA had a partly different open reading frame encoding a 50.2 kDa protein (MUTYHbeta). An in vitro translation of type b and c mRNAs produced a 50 kDa MUTYHalpha and 47 kDa MUTYHbeta, respectively. MUTYHalpha and MUTYHbeta were detected in wild-type embryonic stem (ES) cells or thymocytes prepared from wild-type mice, but neither MUTYH-null ES cells nor thymocytes prepared from MUTYH-null mice. Both MUTYHalpha and MUTYHbeta were mainly localized in the nuclei and some in mitochondria in wild-type ES cells. Recombinant MUTYHalpha and beta were expressed as fusion proteins with thioredoxin in Escherichia coli, but only MUTYHalpha was partly soluble and thus could be purified. Recombinant MUTYHalpha possessed DNA glycosylase activities to excise adenine opposite 8-oxoguanine and guanine but not AP lyase activity.