DNA replication, transcription and translation

In very general terms, what does a chromosome contain?

  • Information, genetic information to carry out the characteristics of life -- precise self replication, ability to exchange energy with the environment, etc.

In very general terms, what are the two related functions of DNA?

  • Information storage
  • DNA replication
  • Information transfer
  • DNA transcribed into RNA
  • DNA's function in information transfer

What is the Central Dogma associated with information storage and retrieval?

  • Central Dogma:
  • DNA-->RNA-->unfolded protein-->native, folded protein

What are the three processes of the central dogma? How does DNA function as an information molecule?

  • replication, DNA --> DNA
  • transcription, DNA --> RNA
  • translation, RNA --> unfolded protein --> folded protein

In terms of molecular conformation, what occurs through the central dogma?

  • Translation of linear information, a sequence of nucleotides, into 3-D information, the structure of a protein.

What are the differences between DNA and RNA?

  • base composition: RNA = AGCU, DNA = AGCT
  • carbohydrate: RNA = ribose, DNA = deoxyribose
  • structure: RNA = single stranded, DNA = double helix


  • usually single stranded
  • linear polymer of ribonucleotides.
  • Some secondary and tertiary structure but often ill-defined.

What are the different types of RNA? What are the functions of the different types of RNA?

  • messenger RNA = mRNA, information transfer
  • transfer RNA = tRNA, information transfer
  • ribosomal RNA = rRNA, structural
  • small nuclear RNA = snRNA, ribozymes, RNA processing.

What is replication?

  • Transfer of genetic information from one generation to the next.
  • DNA-directed DNA synthesis: replication of the genome.
  • What is the structural basis for the precise duplication of the genome?
  • The Watson-Crick structure of DNA: the strands are complementary, the nucleotide sequence in one automatically specifies the other.
  • The enzyme, DNA polymerase III, is very accurate: it has proof reading capabilities.
  • Is replication conservative or semi-conservative? What does that mean?
  • Is the parental genome of double stranded DNA fully conserved in the parental cell or is it split equally (semi-conserved) between two daughter cells?
  • Replication is semi-conservative.

What is the evidence for semi-conservative replication?

  • Classical experiments of Meselson and Stahl. Label DNA with *heavy isotope* N15 and allow replication in light N14: distinguish heavy, light and hybrid DNA by centrifugation.
  • Results: after 1 generation, each genome contains a hybrid N15-N14 DNA; after 2 generations, there are 2 hybrid and 2 light (N14-N14) genomes.
  • Each strand of DNA serves as a template for the synthesis of its complement.
  • The strands separate and each is used as a template for the synthesis of a daughter strand.
  • The two new double helices each contain half the parental DNA.
  • This process produces a replication fork

Is replication uni-directional or bi-directional?

  • Bi-directional
  • Two replication forks proceeding from the origin.

What is the major replication enzyme?

  • DNA polymerase III, a DNA-directed DNA polymerase
  • Synthesis is 5'-->3'
  • Substrates are deoxynucleoside triphosphates (to make deoxyribonucleic acid)
  • Proof reading , errors removed by 3'--5' exonuclease
  • Processivity is very high (the ability of the enzyme to replicate a large tract of DNA before *falling* off)
  • Replication requires DNA unwinding by enzymes termed helicases: these enzymes unwind the DNA helix before the replication fork and wind it up again afterwards.
  • There are large numbers of different enzymes and proteins involved at the replication fork in the replisome.
  • DNA damage by UV radiation or chemicals is repaired by other DNA polymerases. UV-damage results in adjacent T residues in one strand becoming covaletly linked to each other, producing a thymine dimer. This causes the double helix to become distorted -- kinky. Xeroderma pigmentosa is a genetic disorder in which patients cannot carry out UV-radiation repair. They are very prone to skin cancer from an early age.

What is Transcription?

  • Copying a gene as RNA
  • DNA-directed RNA synthesis from a gene

What is a gene?

  • There is no good definition of a gene!
  • A sequence of DNA that is transcribed from specific start to specific stop base sequences.
  • Beadle and Tatum, working with the eukaryote mold Neurospora crassa, concluded that one gene codes for one protein.

But what about genes that code for RNA's like rRNA and tRNA?

  • A gene is a sequence of DNA that is transcribed into a single RNA as defined by specific start and stop sequences of bases.
  • Note the circularity of the argument!
  • But the single RNA may be polycistronic!

What does that mean?

  • A cistron is synonymous with a gene.
  • A polycistronic RNA results from the transcription of an operon.


What's an operon?

  • A genetic unit containing several genes with related functions: the bacterial operon for lactose (milk sugar) metabolism contains 3 genes coding for 3 different proteins.
  • An operon is transcribed as a single unit, a polycistronic messenger RNA (mRNA) that codes for more than one gene product.

Name 4 types of RNA. What are their functions?

  • mRNA, messenger RNA that is translated into protein
  • rRNA, ribosomal RNA that, together with ribosomal proteins, forms a structural scaffold for the translation of mRNA, the ribosome
  • tRNA, transfer RNA, a specific carrier of amino acids
  • snRNA, small nuclear RNA involved in processing of mRNA in the nucleus

What is the major transcription enzyme?

  • RNA polymerase, a DNA-directed RNA polymerase
  • RNA synthesis is 5'-->3'
  • substrates are ribonucleoside triphosphates ( to make ribonucleic acid)
  • begins at the promoter, 5' end of the gene
  • processivity is very high, proceeds to 3' end of gene without stopping or falling off the gene
  • proof reading by precise Watson-Crick base pairing, A=U and G=C
  • Regulation of transcription of a gene is at the 5'-end of the gene at region(s) termed operators
  • Transcription of some genes is constitutive = housekeeping genes
  • Transcription of other genes is in response to a stimulus = inducible genes

What are exons and introns?

  • exons are coding regions, and
  • introns are non-coding regions of the mRNA transcript
  • exons and introns are found in most, but not all, eukaryote genes
  • introns have to be spliced out before the mRNA is translated
  • splicing is by snRNA's acting as enzymes, or ribozymes, an example of the catalytic function of RNA


  • Synthesis of a linear polymer of amino acids from a linear polymer of nucleotide

Where does it occur?


  • On the ribosome, a rRNA-protein complex that provides:
  • a scaffold for mRNA
  • sites for the docking of tRNA charged with a specific amino acid
  • an enzyme for peptide bond synthesis between amino acids
  • an enzyme for translocation of the mRNA through the ribosome

What is the function of tRNA?

  • Carrier of a specific amino acid during translation

What is the structure of tRNA?

  • secondary structure has some base-pairing --> cloverleaf
  • information transfer at the anti-codon loop, complementary to the codon
  • note the importance of H-bonds in the genetic code
  • tertiary structure is L-shaped which places the amino acid far from the codon-anticodon site
  • degeneracy of the code produces wobble

What is the genetic code?

  • A sequence of 3 nucleotides forms a codon
  • unambiguous, each codon specifies an amino acid, or start, or stop
  • degenerate, some amino acids have multiple codons
  • 2-letters often sufficient, specifiy hydrophobic and hydrophillic amino acids
  • What is the enzyme that charges tRNA with an amino acid?
  • An aminoacyl-tRNA synthetase
  • it has proof reading capabilities through the precise fit of amino acid and tRNA
  • energy provided by ATP: energy for the formation of aminoacyl-tRNA and for proof reading
  • there are at least 20 synthetases, isoaccepting for the tRNA's coding for a single amino acid

What is the mechanism of translation?

  • mRNA forms a large complex with the ribosome and protein factors
  • together they guide in the correct aminoacyl-tRNA
  • correct amino acid specified by codon-anticodon base pairing (H-bonds)
  • protein factors have proof reading capability--energy provided by GTP
  • an enzyme catalyzes polymerization of two amino acids, peptide (amide)bond formation between two amino acids
  • an enzyme catalyzes movement of mRNA through the polymerization site: energy provided by GTP
  • mRNA translated from 5'--> 3', same direction as it is synthesized
  • Reprise:
  • Flow of information: central dogma
  • DNA--> RNA-->linear amino acid sequence --> 3D-conformation of protein
  • But some viruses have only RNA as their genome: no DNA.

How do they carry out information transfer? How do they get around the unidirectional flow of information in the central dogma?

  • Use an enzyme called reverse transcriptase to transcribe RNA into DNA.
  • Example: HIV, a retrovirus
  • Then, use central dogma.
  • For HIV:
  • RNA-->DNA--> mRNA --> linear amino acid sequence --> 3D-conformation of protein.

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