第7章rRNA與tRNA的加工

1、RNA Processing Very few RNA molecules are transcribed directly into the final mature RNA. Most newly transcribed RNA molecules (primary transcripts) undergo various alterations to yield the mature product RNA processing 是對(duì)初生轉(zhuǎn)錄物進(jìn)行加工是對(duì)初生轉(zhuǎn)錄物進(jìn)行加工修飾的總稱。修飾的總稱。RNA processingRomoval of nucleotidesaddition o

2、f nucleotides to the 5- or 3- endsmodification of certain nucleotides(1) 外切、內(nèi)切核酸酶對(duì)堿基的切割外切、內(nèi)切核酸酶對(duì)堿基的切割(2) 向初生轉(zhuǎn)錄物的向初生轉(zhuǎn)錄物的3端和端和5端添加端添加核苷酸核苷酸(3) 對(duì)某些特殊的核苷酸或糖苷進(jìn)行修飾對(duì)某些特殊的核苷酸或糖苷進(jìn)行修飾Add a methyl group to 2-OH of ribose in mRNA (A) and rRNAExtensive changes of bases in tRNARNPsRibonucleoproteins = RNA protein

3、 complexes3-D structuretRNA PROCESSING, RNase P AND RIBOZYMES 1. tRNA processing in prokaryotes2. tRNA processing in eukaryotes3. RNase P4. Ribozymes1. tRNA processing in prokaryotesMature tRNAs are generated by processing longer pre-tRNA transcripts, which involves 1.specific exo- and endo- nucleol

4、ytic cleavage by RNases D, E, F and P (general) followed by 2.base modifications which are unique to each particular tRNA type.大腸桿菌tRNA操縱子中包含tRNA的編碼區(qū)域；還包括多至7種tRNA基因的操縱子1. 轉(zhuǎn)錄物前體折疊形成特殊的莖環(huán)結(jié)構(gòu)后，內(nèi)切核酸酶（內(nèi)切核酸酶（RNase E，F(xiàn)）在3端切割一個(gè)側(cè)翼序列，在前體的3端留下額外的9核苷酸；2. 外切酶RNase D依次切去3端7個(gè)核苷酸；3. RNase P內(nèi)切產(chǎn)生成熟的5端；4. RNase D切去剩余的2

5、個(gè)核苷酸，產(chǎn)生成熟的3端。原核生物原核生物tRNA的加工過(guò)程的加工過(guò)程大腸桿菌中前大腸桿菌中前tRNA 的加工過(guò)程的加工過(guò)程2. tRNA processing in eukaryotesThe pre-tRNA is synthesized with a 1. 16 nt 5-leader, 2. a 14 nt intron and 3. two extra 3-nucleotides.leaderintrontwo extra真核生物前tRNA的結(jié)構(gòu)與加工過(guò)程示意圖1. Primary transcripts forms secondary structures recognized b

6、y endonucleases 2. 5 leader and 3 extra nucleotide removal3. tRNA nucleotidyl transferase (核酸轉(zhuǎn)核酸轉(zhuǎn)移酶移酶)adds 5-CCA-3 to the 3-end to generate the mature 3-end4. Intron removal真核生物前真核生物前tRNA的加工過(guò)程的加工過(guò)程原核真核原核真核tRNA加工過(guò)程的不同點(diǎn)加工過(guò)程的不同點(diǎn) 真核生物前tRNA中含有內(nèi)含子； 真核生物的3端CCA是由核苷酸轉(zhuǎn)移酶添加上的。3. RNase P Ribonuclease P (RNase P

7、) is an enzyme involved in tRNA processing that removes the 5 leader sequences from tRNA precursors RNase P (1)RNase P 由一由一RNA分子和蛋白質(zhì)分子組成的分子和蛋白質(zhì)分子組成的內(nèi)切酶，是一類比較簡(jiǎn)單的內(nèi)切酶，是一類比較簡(jiǎn)單的RNP；作用：使前作用：使前tRNA分子產(chǎn)生成熟的分子產(chǎn)生成熟的5端；端；分布：真核原核都有發(fā)現(xiàn)分布：真核原核都有發(fā)現(xiàn)1. EU：位于核內(nèi)，是核內(nèi)小分子：位于核內(nèi)，是核內(nèi)小分子RNP，snRNP2. Pro: 377nt+13.7KD的蛋白質(zhì)的蛋白質(zhì)RNa

8、se P (2) RNA component can catalyze pre-tRNA in vitro in the absence of protein. Thus RNase P RNA is a catalytic RNA, or ribozyme.4. Ribozyme 核酶是一類能夠催化特定的生化反應(yīng)的核酶是一類能夠催化特定的生化反應(yīng)的RNA分子。分子。 是核糖核酸和酶的是核糖核酸和酶的2個(gè)詞的縮寫(xiě)個(gè)詞的縮寫(xiě) RNase P RNA is a ribozyme. RNase P RNA from bacteria is more catalytically active in v

9、itro than those from eukaryotic and archaebacterial （古（古細(xì)菌）細(xì)菌）cells. All RNase P RNAs share common sequences and structures. 核酶的錘頭結(jié)構(gòu) 該酶包括三個(gè)莖環(huán)區(qū)，其間有一個(gè)11-13個(gè)保守核苷酸構(gòu)成的催化中心。 N代表該位置上可以為任意堿基， 箭頭代表切割部位， H代表除了G以外的核苷酸。 在類病毒中，莖III的頂部并沒(méi)有被環(huán)連接起來(lái)RibozymeSelf-splicing introns: the intervening RNA that catalyze the s

10、plicing of themselves from their precursor RNA, and the joining of the exon sequences1.Group I introns, such as Tetrahymena（四膜蟲(chóng)）（四膜蟲(chóng)） intron2.Group II introns. Ribozyme 的用處的用處可以人工合成核酶，作診斷治療：可以人工合成核酶，作診斷治療：1.correcting mutant mRNA in human cells2. inhibiting unwanted gene expressionKill cancer cellsP

11、revent virus replication核酶的生物學(xué)意義核酶的生物學(xué)意義 核酶的發(fā)現(xiàn)使我們對(duì)RNA的重要功能又有了新的認(rèn)識(shí)。 核酶是繼逆轉(zhuǎn)錄現(xiàn)象之后對(duì)中心法則的又一個(gè)重要修正，說(shuō)明RNA既是遺傳物質(zhì)又是酶。 核酶的發(fā)現(xiàn)為生命起源的研究提供了新思路。Splicing of Group I and II introns Introns in fungal mitochondria, plastids, Tetrahymena(四膜蟲(chóng)四膜蟲(chóng)) pre-rRNA Group I Self-splicing Initiate splicing with a G nucleotide Group

12、II self-splicing Initiate splicing with an internal AtRNA PROCESSING1. rRNA processing in prokaryotes2. rRNA processing in eukaryotesrRNA processing in prokaryotes 在大腸桿菌中，rRNA合成由七種分散在基因組中的RNA操縱子完成； 每一個(gè)操縱子都包括5S rRNA,16S rRNA和23S rRNA各一個(gè)，及1-4個(gè)tRNA的編碼序列； 初生轉(zhuǎn)錄物沉降系數(shù)為30S，加工形成成熟的rRNA和tRNA； 對(duì)酶缺陷型的研究表明大腸桿菌的M5,M16,M23,RNA酶III都參與了rRNA的加工過(guò)程大腸桿菌前rRNA加工過(guò)程1. 轉(zhuǎn)錄物通過(guò)堿基配對(duì)形成莖環(huán)結(jié)構(gòu)；2. RNA酶III剪切并釋放5S，16S，23S的前體分子；3. RNA酶M5，M16，M23分別在每一前體分子的5和3端進(jìn)一步剪切，在第二部剪切后釋放出成熟的全長(zhǎng)rRNA大腸桿菌前rRNA加工過(guò)程rRNA processing in eukaryotes真核生物的rRNA由RNAP I轉(zhuǎn)錄合成；rRNA基因串聯(lián)重復(fù)排列，不同生物的rRNA大小不一；酵母7000nt，哺乳動(dòng)物：13500nt前體包括5.8S，