生物化学资料：Biochemistry(01) - edit

1、Biochemistry,SHARKHEUIRO,1,Modification of RNA,15:27:53,Catalogue,TOPIC,Posttranscriptional Modification Of RNA,01,Ribosomal RNA,02,Transfer RNA,03,Eukaryotic mRNA,04,Comparison Of RNA Processing Between Prokaryotic cell & Eukaryotic cell,2,Modification of RNA,15:27:53,ONE,Ribosomal RNA,Post-transcr

2、iptional modification in E.coli,1. E.coli totally has seven operons , which are dispersed in the genome. Each operon consists of 16S rRNA ，23S rRNA ，5S rRNA and 1-4 tRNA genes,2. The initial transcript has a sedimentation coefficient of 30s (6000 nt) and is normally quite short-lived,operon,3,Modifi

3、cation of RNA,15:27:53,1) Following or during the primary transcription, the RNA folds up into a number of stem-loop structures by base pairing between complementary sequences,The formation of this secondary structure of stems and loops allows some proteins to bind to form a RNP（ribonucleoprotein） c

4、omplex（核糖核蛋白复合体） which remain attached to the RNA,Processing of rRNA,4,Modification of RNA,15:27:53,Processing of rRNA,2) After the binding of proteins, methylation（甲基化） takes place. It is cleavaged by RNase ,and release pre-16S ,pre-23S, pre-5S rRNA molecules,5,Modification of RNA,15:27:53,Processi

5、ng of rRNA,3）Under the action of RNA-ase M5, M16, M23, each of the two ends of the precursor RNA molecule is further cut, and then a mature RNA molecule is released,6,Modification of RNA,15:27:53,Processing of rRNA,1) Eukaryotes have four types of rRNA,respectively 28S , 18S ,5.8S and 5SrRNA 2) 45S

6、rRNA transcript is produced during transcription, which is then processed into 18S, 5.8S and 28S rRNA. These rRNA transcription units are transcribed by RNA pol I. 3) 5S rRNA genes are transcribed by RNA polymerase III,7,Modification of RNA,15:27:53,Processing of rRNA,8,Modification of RNA,15:27:53,

7、Processing of rRNA,9,Modification of RNA,1) Remove the 5 end of the leader sequence , and 45S primary transcript turns into the intermediate 41S 2) The18S fragments are firstly cut from the intermediate 41S, producing respectively 20S (containing 18S rRNA fragment) and 32S 3）Hydrogen bonds between t

8、he 5.8S and 28S in the 32S intermediate are formed, and they form a hairpin structure 4） By RNase cleavage, a mature rRNA is formed ultimately,15:27:53,Processing of rRNA,10,Modification of RNA,15:27:53,TWO,Transfer RNA,1、Primary structure : single strand of RNA , about 73-93nt nucleotides 2、seconda

9、ry structure : The cloverleaf structure , the base paring of various regions form four stems (arms) and three loops,11,Modification of RNA,15:27:53,Processing of tRNA,12,Modification of RNA,15:27:53,Modification of tRNA Tyr in E.coli,Tyr(酪氨酸,反密码子GAU,prokaryotic,13,Modification of RNA,15:27:53,The pr

10、ecursor transcripts are folded to form a characteristic stem-loop structure Rnase F endonuclease excises the flanking sequence at the 3 end . Thus an additional 9 nucleotides are left at the 3 end. Rnase D exonuclease excises 7 nucleotides from the 3 end Rnase P endonuclease excises the leader seque

11、nce of 5end to produce the mature 5 end Rnase D exonuclease continues to remove the remaining two nucleotides of 3 end, to produce the mature 3 end CCA-OH tRNA goes through a series of base modification, and finally mature tRNA molecules are formed,14,Modification of RNA,15:27:53,15,endonuclease,RNA

12、aseP,Leader seq,1. Excision,ligase,Modification of RNA,15:27:53,16,tRNA nucleotidyl transferase,ATP,ADP,2. Tailing,Modification of RNA,15:27:53,17,1） A(G) Am (Gm,3. Base modification,Modification of RNA,2） U DHU,3） U T U,4） A I,15:27:53,THREE,Messenger RNA,Capping of mRNA Polyadenylation RNA Splicin

13、g Alternative Splicing RNA Editing,18,Modification of RNA,15:27:53,prokaryotic,eukaryotic,prokaryotic mRNA is generally identical to its primary transcript,eukaryotic mRNA is extensively modified both co- and posttranscriptionally,19,Modification of RNA,15:27:53,7-methylguanosine,5 capping,The cap i

14、s a 7-methylguanosine attached to the 5-terminal end of the mRNA through an unusual 5-5 triphosphate linkage,20,Modification of RNA,15:27:53,5 pppGp,Formation of the 5 cap,21,Modification of RNA,15:27:53,Modification of RNA,m7GPPPN,m7GPPPNm,m7GPPPNmNm,Methylate riboses of the two nucleotide in 5end,

15、Methylate the ribose of the first nucleotide in 5end,species of cap,22,Modification of RNA,15:27:53,Modification of RNA,single cell,most eukaryotic cell except for single cells,10%-15,23,Modification of RNA,15:27:53,Modification of RNA,24,Modification of RNA,15:27:53,Modification of RNA,Polyadenylat

16、ion,A chain of 40-250 adenine nucleotides attached to the 3-end,25,Modification of RNA,15:27:53,Modification of RNA,26,Modification of RNA,CPSF:cleavage and polyadenylation specificity factor 聚腺苷酸化特异因子 CF:cleavage factor断裂因子,CstF:cleavage stimulatory factor断裂激动因子 PAP:polyadenylate polymerase 多聚腺苷酸聚合

17、酶 PBP:poly(A)结合蛋白,15:27:53,Functions of polyadenylation,27,Modification of RNA,15:27:54,INTRON,The word intron is derived from the term intragenic region, i.e. a region inside a gene,An experiment that proves the pre-mRNA was spliced for real,DNARNA hybridization,28,Modification of RNA,15:27:54,INTR

18、ON,The word intron is derived from the term intragenic region, i.e. a region inside a gene,Introns are divided into two types according to the process of its splicing,Self-splicing intron,Intron,Spliceosomal intron,Group I,Group II,mainly exists in organelle,GU-AG Intron,AU-AC Intron,mostly exists i

19、n nucleus,29,Modification of RNA,15:27:54,SPLICING,The process of removing introns and joining exons is call splicing . Spliceosome accomplishes these tasks,Removal of introns and intervening sequences , which do not code for protein,The remaining coding sequences , the exons , are joined together t

20、o form the mature mRNA,30,Modification of RNA,15:27:54,Self-Splicing,Self-splicing occurs for rare introns that form a ribozyme, performing the functions of the spliceosome by RNA alone,Group I: 3-OH of GMP attack the phosphodiester bond at 5 as a nucleophilic group of intron. freed 3-OH of upstream

21、 exon attack the phosphodiester bond at the 3 of intron,Group II: 2-OH of AMP attack the phosphodiester bond at 5 as a nucleophilic group of intron. freed 3-OH of upstream exon attack the phosphodiester bond at the 3 of intron,31,Modification of RNA,15:27:54,The splice donor site includes an almost

22、invariant sequence GU at the 5 end of the intron. The splice acceptor site at the 3 end of the intron terminates the intron with an almost invariant AG sequence,GU-AG Intron Splicing,The branch site is close to 3 splice site , upstream of polypyrimidine tract(多聚嘧啶区,32,Modification of RNA,15:27:54,Th

23、e branch site is close to 3 splice site , upstream of polypyrimidine tract,GU-AG Intron Splicing,2-OH of branch site A attacks the phosphate at 5-end of intron , forming 25 phosphodiester bond and creating a lariat,Freed 3-OH of exon 1 attacks 5-phosphate at splice acceptor site , forming a phodiest

24、er bond,Excised intron released , and then degraded,33,Modification of RNA,15:27:54,SPLICEOSOME,Catalyzes pre-mRNA splicing in nucleus. Composed of snRNAs and snRNPs,34,Modification of RNA,15:27:54,snRNPs,Uracil-rich snRNAs , mainly including U1 , U2 , U4 , U5 and U6 , along with multiple proteins f

25、orm small nuclear ribonucleoprotein particles that mediate splicing,U1 snRNA With a conserved sequence: 5-UACUUAC-3 , which is complementary with the board sequence at 5-end of intron(donor splicing junction,35,Modification of RNA,15:27:54,snRNPs,Uracil-rich snRNAs , mainly including U1 , U2 , U4 ,

26、U5 and U6 , along with multiple proteins form small nuclear ribonucleoprotein particles that mediate splicing,U6 snRNA can pair with either U4 or U2 U6 pairs with the 5 splice site,36,Modification of RNA,15:27:54,snRNPs,Uracil-rich snRNAs , mainly including U1 , U2 , U4 , U5 and U6 , along with mult

27、iple proteins form small nuclear ribonucleoprotein particles that mediate splicing,37,Modification of RNA,15:27:54,Concrete Mechanism Of GU-AG Intron Splicing,Splicing cycle,38,Modification of RNA,15:27:54,A genetic mutation that inserts, deletes or changes a number of nucleotides in the specific site at which splicing of an intron takes place during the processing of precursor messenger RNA into mature messenger RNA,Splice Site Mutations,Can lead to improper splicing and the production of aberrant proteins,39,Modification of RNA,15:27:54,Alternative Splicing,Alternati