RNA synthesis

RNA types
1. 3 main types of RNAs:
  1. mRNA: messenger RNA, codes for proteins.
    1. 3-5% total RNA.
  2. tRNA: transfer RNA, participates to translation.
  3. rRNA: ribosomal RNA, major constituent of ribosomes.
    1. There are 4 main rRNAs.
  4. Noncoding RNAs serve structural and enzymatic functions
Transcription
1. the process of RNA synthesis from a DNA template.
2. Genes can be on either strand of the DNA
  1. But rarely overlap
3. Transcription creates supercoiling
  1. Topoisomerases release supercoils to allow progression
4. Transcription for a gene starts at its promoter
  1. Sequences in the DNA tell RNA polymerases where to start
  2. Activators are also required to initiate transcription
    1. General transcription factors versus regulatory transcription factors
    2. Many of these proteins bind directly to the DNA
RNA structure
1. Primary structure of RNA
  1. RNA = Ribonucleic Acid
  2. 4 major differences between DNA and RNA:
    1. Contains ribose (instead of deoxy-ribose)
    2. Contains U instead of T
    3. RNA is unstable
    4. Synthesized as a single strand
2. Secondary structure of RNA
  1. Pairing between bases: A = U, G = C
  2. Also can rarely have non Watson-Crick pairing, such as G = U.
  3. Formation of stem-loop secondary structures by intramolecular base pairing
3. Secondary and tertiary structure of RNA
  1. Tertiary structure is the folding of a molecule in three dimensions.
Transcription enzymes:RNA polymerases
1. RNA polymerases are enzymes performing RNA synthesis. There are 3 types of RNA polymerases in eukaryotes:
  1. RNA polymerase I
    1. ribosomal RNA
  2. RNA polymerase II
    1. protein-encoding genes
  3. RNA polymerase III
    1. tRNA, small nuclear RNA, and 5S rRNA
2. Transcription requires RNA polymerase
  1. Different from DNA synthesis:
    1. No primer needed to get started
    2. Only one strand of DNA is used as the template
    3. Transcript does not remain bound to template
    4. Higher error rate
    5. Multiple RNA Pol bind on the same gene
RNA processing
1. splicing of introns
  1. eliminates non coding regions of mRNAs to generate mature mRNA for protein synthesis.
2. capping of 5’ end
  1. necessary for stability, binding of mRNA to ribosomes and initiation of translation.
3. polyadenylation of 3’ end
  1. part of the termination process (addition of poly-A tail). Determine mRNA stability, helps mRNA nuclear export and translation.
4. Splicing
  1. Specific to eukaryotes transcription.
  2. Introns are present in the DNA and pre-mRNA, not in mRNA
  3. Some mRNA can be spliced in more than one way
    1. alternative splicing
  4. Splice donor and acceptor sites
    1. 2’ OH of branch site A attacks phosphodiester bond on donor site G.
      1. Cleavage at the donor site. Formation of lariat.
        3’ OH of donor site G attacks phosphodiester bond on acceptor site G freeing the lariat.
        Lariat is degraded
  5. mRNA splicing is performed by the spliceosome
    1. Spliceosome: nuclear complex made of about 150 proteins + 5 RNAs.
      1. Three functions
        recognize 5’ donor and branch sites
        bring sites together
        catalyzes RNA cleavage
    2. SnRNP = SnRNA + proteins
      1. snRNAs are small nuclear RNA: U1, U2, U4, U5 and U6.
5. Polyadenylation
  1. CstF: Cleavage stimulating factor
  2. CTD: Pol II C-terminal domain
  3. CPSF: Cleavage and Polyadenylation specific factor
  4. PAP: polyadenylate polymerase

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RNA synthesis

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