Structural studies on the RNA-recognition motif of NELF E, a cellular negative transcription elongation factor involved in the regulation of HIV transcription
Structural studies on the RNA-recognition motif of NELF E, a cellular negative transcription elongation factor involved in the regulation of HIV transcription
复制标题
DOI：
10.1042/bj20060421
复制
发表时间：
2006-12-15
影响因子：
4.1
通讯作者：
Woehrl, Birgitta M.
中科院分区：
文献类型：
--
作者：
Rao, Jampani N.;Neumann, Liane;Woehrl, Birgitta M.
HIV RNA在TAR（反式激活反应元件）转录的延长受到正性和负性因素的高度调节。细胞负性转录延伸因子NELF（negative elongation factor）被认为通过结合到由细胞RNA聚合酶II在病毒长末端重复序列处合成的病毒TAR RNA的茎部而参与HIV-1（HIV 1型）的转录调节。NELF是由NELF A、B、C或剪接变体D和E组成的异源四聚体蛋白。在本研究中，我们确定了RNA结合亚基NELF E的RRM（RNA识别基序）的溶液结构，并研究了其与病毒TAR RNA的相互作用。我们的结果表明，单独表达的重组NELF E RRM具有采用β α β α β折叠的α-螺旋和P-链元件，并且能够结合TAR RNA。荧光平衡滴定与荧光标记的双链和单链寡核糖核苷酸代表的TAR RNA茎意味着NELF E RRM结合的单链TAR RNA的Kd值在低微摩尔范围。
The elongation of transcription of HIV RNA at the TAR (transactivation-response element) is highly regulated by positive and negative factors. The cellular negative transcription elongation factor NELF (negative elongation factor) was suggested to be involved in transcriptional regulation of HIV-1 (HIV type 1) by binding to the stem of the viral TAR RNA which is synthesized by cellular RNA polymerase II at the viral long terminal repeat. NELF is a heterotetrameric protein consisting of NELF A, B, C or the splice variant D, and E. In the present study, we determined the solution structure of the RRM (RNA-recognition motif) of the RNA-binding subunit NELF E and studied its interaction with the viral TAR RNA. Our results show that the separately expressed recombinant NELF E RRM has a-helical and P-strand elements adopting a beta alpha beta beta alpha beta fold and is able to bind to TAR RNA. Fluorescence equilibrium titrations with fluorescently labelled double- and single-stranded oligoribonucleotides representing the TAR RNA stem imply that NELF E RRM binds to the single-stranded TAR RNAs with K-d values in the low-micromolar range.