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slide16 - #16 RNA processing (II) 1. 2. 3. 4. 5. 6. 7. tRNA...

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#16 RNA processing (II) 1. tRNA and rRNA processing 2. RNA enzymes-ribozymes 3. RNA trans-splicing 4. RNA editing 5. RNA degradation 6. miRNA, siRNA, and RNAi 7. RNA and evolution
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1. tRNA and rRNA processing 1. tRNAs and rRNAs of both eukaryotes and prokaryotes are often synthesized as longer precursors, and need to be processed to become the final products. 2. rRNA and tRNAs are processed by intron splicing, cleavage (cut in the middle without religation), and trimming (cut from ends). Some rRNA/tRNA precursors can be cut to produce several rRNA as well as tRNAs. Many different RNases are involved.
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2. Ribozyme An E. coli RNase called RNase P is involved in tRNA processing. RNase P is composed of a RNA called M1 RNA and a 14kD protein. It was found in 1983 by Thomas Cech (who also first found self-splicing in 1982) that M1 RNA cuts the pre-RNA ( Pre-tRNA tyr ). The M1 RNA and other catalytic RNAs were later named as ribozymes (RNA enzymes).
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Ribozyme catalysis Result : (1) pTyr (pre-tRNA Tyr ) was cut by M1 RNA, but P4.5 (a pre- rRNA) was not cut, demonstrating specificity (2) M1 need more Mg than the RNase P holoenzyme, indicating that although RNA alone is catalytically active, the protein component changes the properties of the enzyme. Experiment : 32 P-label pre-tRNA (and a control pre-rRNA), incubate with M1 RNA, run a gel to see if pre-tRNA is cut by M1. M1+P M1
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Self-cleavage ribozymes (a) Secondary structure of the HDV (Hepatitis delta virus) ribozyme ; the cytosine ( C75 ) is essential to catalysis. The 5'end of the HDV ribozyme RNA marks the cleavage site (between U-C). (b) 3D structure of the HDV ribozyme. The C75 folds to become close to the cleavage site and the 5'end of the ribozyme. (c) Chemistry of the catalysis by C75, in which the protonated C75 donates a proton to catalyze the cleave. ( Nature 2002, 418:222)
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3. Trans-splicing The pre-mRNA splicing discussed before involved cut/religation of fragments within the same RNA molecule, it is called cis-splicing. It was found that, in worm trypanosomes , fragments of different pre-mRNAs can be spliced together, it is called trans-splicing. Trans-splicing involves transesterification reaction without lariat . The previously discussed pre-tRNA cleavage by M1 RNA of RNase P of E. coli can also be called trans-splicing (without re-ligation). (RNA1) (RNA2)
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4. RNA editing Deaminase RNA editing changes individual nucleotides in the pre-RNA to produce mature RNA. Many organellar RNAs are edited. (deamination) (U deletion/insertion) RNA of nuclear genes is rarely edited. RNA editing can change C to U by deaminase, or U deletion/insertion by gRNA (guide RNA), endonuclease, exonuclease, TUTase (terminal U transferase), and RNA ligase.
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ApoB-100 binds to LDL (low density lipoprotein) particle and LDL receptor on the cell surface, resulting in receptor-mediated endocytosis and hydrolysis of LDL and cholesterol in lysosomes of liver cells. A higher ApoB-100 level indicates higher LDL level and increased chances of heart attack and strokes. e.g.RNA editing and cholesterol metabolism
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This note was uploaded on 02/20/2009 for the course LS 3 taught by Professor Lin during the Spring '06 term at UCLA.

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slide16 - #16 RNA processing (II) 1. 2. 3. 4. 5. 6. 7. tRNA...

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