Biochemistry

• Gene Expression (Protein Biosynthesis)

  Transcription and Translation
• Transcription


  DNA-directed RNA synthesis
• The same genetic information is in all 100 trillion cells of any person, but different cells use the same blueprint in different ways
• The control of gene expression occurs by regulating the flow of information from DNA to protein

  
• Gene Expression


  • ATCGGAAGT
  • TAGCCTTCT

  
• Gene Expression


  • 5-ACGTTATGAAGTAACCGGCCA-3
  • 3-TGCAATACTTCATTGGCCGCA-5

  
• Transcription and Translation in Prokaryote and Eukaryotes

  • In a eukaryotic cell, the nuclear envelope separates transcription from translation
  • Extensive RNA processing occurs in the nucleus

  
• RNA


  A nucleic acid that uses a slightly different sugar than DNA and the base uracil (U) in place of thymine (T)
• RNAs vs. DNA


  • RNA is single stranded, not double stranded like DNA
  • RNA is short, only 1 gene long, where DNA is very long and contains many genes
  • RNA uses the sugar ribose instead of deoxyribose in DNA
  • RNA uses the base uracil (U) instead of thymine (T) as in DNA

  
• Recently, a new class of RNA, microRNA, has been shown to regulate gene expression

  
• Transcription


  RNA synthesis catalyzed by RNA polymerase, which opens the DNA strands and hooks together the nucleotides
• Transcription (Synthesis of RNA Transcript)

  1. Initiation

  2. Elongation

  3. Termination

  
• Transcription in Prokaryotes - Initiation

  • RNA polymerase binds a region (nucleotides sequence) of the DNA known as the Promoter, which is not transcribed
  • This promoter region recognized by additional subunit of RNA Polymerase called Sigma (σ) Factor
  • Include: –35 sequence and Pribnow box

  
• Transcription in Prokaryotes - Initiation

  • Local unwinding of DNA caused by RNA polymerase and formation of an open initiation complex

  
• Transcription in Prokaryotes - Elongation

  1. RNA polymerase synthesizes a single strand of RNA against the DNA template strand (anti-sense strand), adding nucleotides to the 3' end of the RNA chain

  2. Transcription proceeds 5- to 3': new bases are added to the free 3' OH group

  3. The raw: ATP, CTP, GTP, and UTP

  
• Transcription in Prokaryotes - Termination

  1. Specific sequences in the DNA signal termination of transcription

  2. When one of these is encountered by the polymerase, the RNA transcript is released from the DNA and the double helix can zip up again

  3. A protein called rho (ρ) factor is required for termination of transcription of some genes in prokaryotes

  
• Antibiotics


  Some antibiotics prevent bacterial cell growth by inhibiting RNA synthesis
• Antibiotics


  • Rifampin (treatment of tuberculosis): Inhibits the initiation of transcription by binding to the β subunit of prokaryotic RNA polymerase make it non functional
  • Dactinomycin (known to biochemists as actinomycin D) binds to the DNA template and interferes with the movement of RNA polymerase along the DNA
• Transcription in Eukaryotes


  • There are three distinct classes of RNA polymerase in the nucleus of eukaryotic cells
  • RNA polymerase I: synthesizes the rRNA
  • RNA polymerase II: synthesizes mRNA (and some ncRNAs)
  • RNA polymerase III synthesizes tRNA in the nucleoplasm

  
• Transcription in Eukaryotes


  • Promoters for genes transcribed by RNA polymerase II include: TATA (or Hogness) box, CAAT box, and GC box
  • They serve as binding sites for proteins called general transcription factors, which, in turn, interact with each other and with RNA polymerase II

  
• Enhancers


  DNA sequences that increase the rate of initiation of transcription by binding specific transcription factors that serve as transcription activators
• Post-Transcriptional Modification (RNA Processing)

  • Most eukaryotic mRNAs aren't ready to be translated into protein directly after being transcribed from DNA
  • RNA processing occur in the nucleus
  • The mature mRNA moves to the cytoplasm for translation
  • Long single Pre-rRNA cleaved by ribonucleases and then trimmed by endonucleases
  • Base and sugar modifications by snoRNA

  
• mRNA Processing


  1. 5' Capping: Protection, Unusual 5'→5' triphosphate linkage made between 5'-terminal end of the mRNA and 7-methylguanosine

  2. Poly A Tail: Protection, stabilizing and facilitate exit, 40–200 adenine nucleotides attached to the 3'-end, not transcribed from the DNA, Added after transcription by the nuclear enzyme, polyadenylate polymerase, using ATP as the substrate

  3. Splicing: Most of the eukaryotic genome consists of non-coding regions called introns, Introns are removed from pre-RNA transcript while it is still in the nucleus, Spliceosome composed of small-nuclear ribonucleoprotein particles (snRNPs), that mediate the process of splicing, consist of (proteins + uracil-rich small nuclear RNAs (snRNA)

  
• After splicing the remaining ends are re-attached, so the final RNA consists of exons only
• Systemic lupus erythematosus, SLE; an often fatal inflammatory disease, results from an autoimmune response in which individuals produce antibodies against their own nuclear proteins such as snRNPs
• Gene Expression (Protein Biosynthesis)

  Transcription and Translation
• Transcription


  DNA-directed RNA synthesis
• The same genetic information is in all 100 trillion cells of any person, but different cells use the same blueprint in different ways
• The control of gene expression occurs by regulating the flow of information from DNA to protein
• Gene Expression


  • ATCGGAAGT
  • TAGCCTTCT

  
• Gene Expression


  • 5-ACGTTATGAAGTAACCGGCCA-3
  • 3-TGCAATACTTCATTGGCCGCA-5

  
• Transcription and Translation in Prokaryote and Eukaryotes

  • In a eukaryotic cell, the nuclear envelope separates transcription from translation
  • Extensive RNA processing occurs in the nucleus

  
• RNA


  A nucleic acid that uses a slightly different sugar than DNA and the base uracil (U) in place of thymine (T)
• RNAs vs. DNA


  • RNA is single stranded, not double stranded like DNA
  • RNA is short, only 1 gene long, where DNA is very long and contains many genes
  • RNA uses the sugar ribose instead of deoxyribose in DNA
  • RNA uses the base uracil (U) instead of thymine (T) as in DNA

  
• Recently, a new class of RNA, microRNA, has been shown to regulate gene expression

  
• Transcription


  RNA synthesis catalyzed by RNA polymerase, which opens the DNA strands and hooks together the nucleotides
• Transcription (Synthesis of RNA Transcript)

  1. Initiation

  2. Elongation

  3. Termination

  
• Transcription in Prokaryotes - Initiation

  • RNA polymerase binds a region (nucleotides sequence) of the DNA known as the Promoter, which is not transcribed
  • This promoter region recognized by additional subunit of RNA Polymerase called Sigma (σ) Factor
  • Include: –35 sequence and Pribnow box

  
• Transcription in Prokaryotes - Initiation

  • Local unwinding of DNA caused by RNA polymerase and formation of an open initiation complex

  
• Transcription in Prokaryotes - Elongation

  1. RNA polymerase synthesizes a single strand of RNA against the DNA template strand (anti-sense strand), adding nucleotides to the 3' end of the RNA chain

  2. Transcription proceeds 5- to 3': new bases are added to the free 3' OH group

  3. The raw: ATP, CTP, GTP, and UTP