A three-dimensional model for the hammerhead ribozyme based on fluorescence measurements.
A three-dimensional model for the hammerhead ribozyme based on fluorescence measurements.
复制标题
基于荧光测量的锤头核酶三维模型。
作者：
T. Tuschl;C. Gohlke;T. Jovin;E. Westhof;F. Eckstein
为了了解RNA锤头核酶的催化功能，三维模型是必不可少的，但既没有晶体也没有溶液结构。利用荧光共振能量转移（Fluorescence resonance energy transfer， FRET）研究溶液中核酶的结构，建立三个组成的沃森-克里克碱基配对螺旋片段的相对空间取向。合成结构体用荧光供体（5-羧基荧光素）和受体（5-羧基四甲基罗丹明）标记在构成核酶分子的链的两端。在螺旋对I和III以及II和III中，受体螺旋的长度分别在5到11和5到9个碱基对之间变化，FRET效率被确定并与一组标记的RNA双链相关。根据向量代数分析预测了FRET效率，作为核酶结构中相对螺旋取向的函数，并与实验值进行了比较。这些数据与核酶的y形排列一致，螺旋I和II靠得很近，螺旋III指向远处。这些取向约束被用于完整核酶的三维结构的分子建模。
For the understanding of the catalytic function of the RNA hammerhead ribozyme, a three-dimensional model is essential but neither a crystal nor a solution structure has been available. Fluorescence resonance energy transfer (FRET) was used to study the structure of the ribozyme in solution in order to establish the relative spatial orientation of the three constituent Watson-Crick base-paired helical segments. Synthetic constructs were labeled with the fluorescence donor (5-carboxyfluorescein) and acceptor (5-carboxytetramethylrhodamine) located at the ends of the strands constituting the ribozyme molecule. The acceptor helix in helix pairs I and III and in II and III was varied in length from 5 to 11 and 5 to 9 base pairs, respectively, and the FRET efficiencies were determined and correlated with a reference set of labeled RNA duplexes. The FRET efficiencies were predicted on the basis of vector algebra analysis, as a function of the relative helical orientations in the ribozyme constructs, and compared with experimental values. The data were consistent with a Y-shaped arrangement of the ribozyme with helices I and II in close proximity and helix III pointing away. These orientational constraints were used for molecular modeling of a three-dimensional structure of the complete ribozyme.
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DOI：
10.1073/pnas.83.24.9373
发表时间：
1986-12-01
影响因子：
11.1
作者：
FREIER, SM;KIERZEK, R;TURNER, DH
通讯作者：
TURNER, DH
影响因子：
2.9
作者：
FEDOR, MJ;UHLENBECK, OC
通讯作者：
UHLENBECK, OC
锤头核酶结构域的核磁共振研究。
影响因子：
5.6
作者：
Heus,HA;Pardi,A
通讯作者：
Pardi,A
自裂解锤头 RNA 茎 I 和 II 的螺旋间几何形状。
影响因子：
2.9
作者：
Gast,FU;Amiri,KM;Hagerman,PJ
通讯作者：
Hagerman,PJ
DOI：
10.1073/pnas.86.20.7706
发表时间：
1989-10-01
影响因子：
11.1
作者：
JAEGER, JA;TURNER, DH;ZUKER, M
通讯作者：
ZUKER, M