The mechanism of pseudouridine synthases from a covalent complex with RNA, and alternate specificity for U2605 versus U2604 between close homologs.
The mechanism of pseudouridine synthases from a covalent complex with RNA, and alternate specificity for U2605 versus U2604 between close homologs.
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DOI：
10.1093/nar/gkt1050
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发表时间：
2014-02
影响因子：
14.9
通讯作者：
Stroud RM
中科院分区：
文献类型：
--
作者：
Czudnochowski N;Ashley GW;Santi DV;Alian A;Finer-Moore J;Stroud RM
RluB 在大肠杆菌 23S rRNA 肽基转移酶中心的茎环中催化 U2605 修饰为假尿苷 (Ψ)。同源物 RluF 对茎 U2604 中的相邻核苷酸具有特异性。 RluB 催化结构域和分离底物茎环之间复合物的 1.3 Å 分辨率晶体结构（其中目标尿苷被 5-氟尿苷 (5-FU) 取代）揭示了异构化目标碱基和酪氨酸 140 之间的共价键。该结构与在 2.5 Å 分辨率下确定的单独催化结构域进行了比较。 RluB 结合的茎环具有与核糖体基本相同的二级结构，在 A2602 处有一个凸起，但 5-FU2605 翻转到活性位点。我们之前表明，RluF 诱导 RNA 移码，将 A2602 移动到茎中，并将其靶标 U2604 翻译到活性位点。氢键网络稳定了 RluB-RNA 中的凸起，但在 RluF 中不保守，因此 RluF 不能稳定凸起。基于酶和异构化 5-FU 之间的共价键，我们提出了与所有实验数据一致的假尿苷形成的迈克尔加成机制。
RluB catalyses the modification of U2605 to pseudouridine (Ψ) in a stem-loop at the peptidyl transferase center of Escherichia coli 23S rRNA. The homolog RluF is specific to the adjacent nucleotide in the stem, U2604. The 1.3 Å resolution crystal structure of the complex between the catalytic domain of RluB and the isolated substrate stem-loop, in which the target uridine is substituted by 5-fluorouridine (5-FU), reveals a covalent bond between the isomerized target base and tyrosine 140. The structure is compared with the catalytic domain alone determined at 2.5 Å resolution. The RluB-bound stem-loop has essentially the same secondary structure as in the ribosome, with a bulge at A2602, but with 5-FU2605 flipped into the active site. We showed earlier that RluF induced a frame-shift of the RNA, moving A2602 into the stem and translating its target, U2604, into the active site. A hydrogen-bonding network stabilizes the bulge in the RluB–RNA but is not conserved in RluF and so RluF cannot stabilize the bulge. On the basis of the covalent bond between enzyme and isomerized 5-FU we propose a Michael addition mechanism for pseudouridine formation that is consistent with all experimental data.
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影响因子：
3.9
作者：
Aravind, L;Koonin, EV
通讯作者：
Koonin, EV
影响因子：
4.5
作者：
Conrad, J;Niu, LH;Ofengand, J
通讯作者：
Ofengand, J
影响因子：
2.9
作者：
Huang, LX;Pookanjanatavip, M;Santi, DV
通讯作者：
Santi, DV
DOI：
10.1073/pnas.0607541103
发表时间：
2006-10-24
影响因子：
11.1
作者：
Berk, Veysel;Zhang, Wen;Cate, Jamie H. Doudna
通讯作者：
Cate, Jamie H. Doudna
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K