A single amino acid substitution in ORF1 dramatically decreases L1 retrotransposition and provides insight into nucleic acid chaperone activity.
A single amino acid substitution in ORF1 dramatically decreases L1 retrotransposition and provides insight into nucleic acid chaperone activity.
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DOI：
10.1093/nar/gkn554
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发表时间：
2008-10
影响因子：
14.9
通讯作者：
Williams MC
中科院分区：
文献类型：
--
作者：
Martin SL;Bushman D;Wang F;Li PW;Walker A;Cummiskey J;Branciforte D;Williams MC
L1是哺乳动物中普遍存在的散布重复序列，其通过自主反转录转座获得高拷贝数。基因组内的单个L1元件在序列和反转录转座活性方面不同。反转录转座需要两个L1编码的蛋白质，ORF1p和ORF2p。嵌合元件被用来映射一个15倍的差异，从小鼠基因组，TFC和TFspa，一个单一的氨基酸取代ORF1p，D159H的两个L1变体之间的反转录转座效率。L1 RNA和蛋白质的稳态水平在这两种元素之间没有显著差异，但在TFC中更早且以更高的频率检测到新的插入，这表明它更有效地将表达的L1中间体转化为新的插入。两个ORF1蛋白进行纯化，并在体外检测其核酸结合和伴侣活性。虽然这两种ORF 1蛋白的RNA和DNA寡核苷酸结合亲和力在很大程度上无法区分，但D159作为核酸伴侣比H159更有效。这些研究结果支持了在L1反转录转座过程中的后期步骤ORF1p核酸分子伴侣活性的要求，扩展了已知对其与核酸的功能相互作用至关重要的ORF1p区域，并增强了对核酸分子伴侣活性的理解。
L1 is a ubiquitous interspersed repeated sequence in mammals that achieved its high copy number by autonomous retrotransposition. Individual L1 elements within a genome differ in sequence and retrotransposition activity. Retrotransposition requires two L1-encoded proteins, ORF1p and ORF2p. Chimeric elements were used to map a 15-fold difference in retrotransposition efficiency between two L1 variants from the mouse genome, TFC and TFspa, to a single amino acid substitution in ORF1p, D159H. The steady-state levels of L1 RNA and protein do not differ significantly between these two elements, yet new insertions are detected earlier and at higher frequency in TFC, indicating that it converts expressed L1 intermediates more effectively into new insertions. The two ORF1 proteins were purified and their nucleic acid binding and chaperone activities were examined in vitro. Although the RNA and DNA oligonucleotide binding affinities of these two ORF1 proteins were largely indistinguishable, D159 was significantly more effective as a nucleic acid chaperone than H159. These findings support a requirement for ORF1p nucleic acid chaperone activity at a late step during L1 retrotransposition, extend the region of ORF1p that is known to be critical for its functional interactions with nucleic acids, and enhance understanding of nucleic acid chaperone activity.
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影响因子：
--
作者：
Martin, Sandra L.
通讯作者：
Martin, Sandra L.
DOI：
10.1073/pnas.0831042100
发表时间：
2003-04-29
影响因子：
11.1
作者：
Brouha, B;Schustak, J;Kazazian, HH
通讯作者：
Kazazian, HH
影响因子：
3.5
作者：
Kulpa, DA;Moran, JV
通讯作者：
Moran, JV
影响因子：
5.6
作者：
Cruceanu, Margareta;Gorelick, Robert J.;Williams, Mark C.
通讯作者：
Williams, Mark C.
影响因子：
5.6
作者：
Martin, SL;Li, JF;Weisz, JA
通讯作者：
Weisz, JA