2013年10月23日,，北京生命科學(xué)研究所葉克窮實(shí)驗(yàn)室在《RNA》雜志發(fā)表題為“Structural and functional analysis of Utp23, a yeast ribosome synthesis factor with degenerate PIN domain”的文章,。
核糖體是所有生物體中合成蛋白質(zhì)的巨大的分子機(jī)器,，在真核生物中由4條RNA和~80個(gè)蛋白質(zhì)構(gòu)成,。真核生物核糖體的裝配也是非常復(fù)雜的過(guò)程,，需要約200個(gè)保守的蛋白質(zhì)因子和很多snoRNA的參與,。Utp23是參與核糖體小亞基早期裝配的保守的蛋白因子,，它含有一個(gè)失去活性的PIN核酸酶結(jié)構(gòu)域和C末端的長(zhǎng)尾巴,。
作者利用蛋白質(zhì)晶體學(xué),，生物化學(xué)和酵母實(shí)驗(yàn)分析了Utp23的結(jié)構(gòu)和功能。他們解析了Utp23 PIN結(jié)構(gòu)域的2.5埃的晶體結(jié)構(gòu),，發(fā)現(xiàn)它除了包含保守的PIN核心結(jié)構(gòu),，還有一些特殊的結(jié)構(gòu)——N末端額外的螺旋，C末端的延伸區(qū)域和一個(gè)鋅指結(jié)構(gòu),。作者還通過(guò)定點(diǎn)突變分析了Utp23的功能位點(diǎn),。他們發(fā)現(xiàn)螺旋1上的多個(gè)保守的堿性氨基酸殘基對(duì)生長(zhǎng)很重要，并和Utp23在體外結(jié)合RNA的活性有關(guān),。他們發(fā)現(xiàn)破壞鋅指結(jié)構(gòu)會(huì)嚴(yán)重的抑制酵母生長(zhǎng),。Utp23的C末端尾巴包含一小段的保守序列，C末端的缺失能破壞Utp23和另一個(gè)核糖體裝配因子snR30 RNA的結(jié)合,，影響Utp23和核糖體前體的結(jié)合,。該工作為理解Utp23在核糖體裝配過(guò)程中的功能提供了重要的結(jié)構(gòu)基礎(chǔ)。
論文的第一作者是我所和北京協(xié)和醫(yī)學(xué)院聯(lián)合培養(yǎng)的博士研究生盧靜。孫夢(mèng)依同學(xué)也參與了此研究工作,。葉克窮博士是本文通訊作者,。此項(xiàng)研究受中國(guó)科技部和北京市科委資助，在北京生命科學(xué)研究所完成,。(生物谷Bioon.com)
生物谷推薦的英文摘要
RNA doi:10.1261/rna.040808.113
Structural and functional analysis of Utp23, a yeast ribosome synthesis factor with degenerate PIN domain
Jing Lu, Mengyi Sun and Keqiong Ye
During synthesis of yeast ribosome, a large complex, called the 90S pre-ribosome or the small subunit processome, is assembled on the nascent precursor rRNA and mediates early processing of 18S rRNA. The Utp23 protein and snR30 H/ACA snoRNA are two conserved components of 90S pre-ribosomes. Utp23 contains a degenerate PIN nuclease domain followed by a long C-terminal tail and associates specifically with snR30. Here, we report the crystal structure of the Utp23 PIN domain at 2.5-? resolution. The structure reveals a conserved core fold of PIN domain with degenerate active site residues, a unique CCHC Zn-finger motif, and two terminal extension elements. Functional sites of Utp23 have been examined with conservation analysis, mutagenesis, and in vivo and in vitro assays. Mutations in each of three cysteine ligands of zinc, although not the histidine ligand, were lethal or strongly inhibitory to yeast growth, indicating that the Zn-finger motif is required for Utp23 structure or function. The N-terminal helix extension harbors many highly conserved basic residues that mostly are critical for growth and in vitro RNA-binding activity of Utp23. Deletion of the C-terminal tail, which contains a short functionally important sequence motif, disrupted the interaction of Utp23 with snR30 and perturbed the pre-ribosomal association of Utp23. Our data establish a structural framework for dissecting Utp23 function in the assembly and dynamics of 90S pre-ribosomes.
