Gene Expression and Regulation

• Rosalind Franklin performed x-ray crystallography of DNA in the 1950s, revealing a regular and repetitive pattern indicative of a double helix
• Edwin Chargoff discovered that the amount of adenine equals the amount of thymine, and the amount of cytosine equals the amount of guanine
• Watson and Crick created the first 3D double helix model of DNA
• DNA is a double-stranded helix with alternating sugar-phosphate backbones and nucleotides in the center
• DNA strands are anti-parallel, with one strand running from 5' to 3' and the other running from 3' to 5'
• DNA is the primary source of heritable information, storing genetic information passed from one generation to the next
• In eukaryotic cells, DNA is linear and found in the nucleus, while in prokaryotic cells, DNA is circular and found in the nucleoid region
• Plasmids are small circular DNA molecules found primarily in prokaryotes, containing advantageous genes that can be manipulated and shared among bacteria
• RNA is single-stranded and contains uracil instead of thymine, while DNA is double-stranded and contains thymine
• Chargoff's rule states that the amount of adenine equals the amount of thymine, and the amount of cytosine equals the amount of guanine
• Purines, such as adenine and guanine, have a double ring structure, while pyrimidines, such as cytosine, uracil, and thymine, have a single ring structure
• Base pairs in DNA are held together by hydrogen bonds, with two hydrogen bonds between adenine and thymine, and three hydrogen bonds between cytosine and guanine
• Three alternative models for DNA replication:

  • Conservative model: Parental strand directs synthesis of an entirely new double-stranded molecule, parental strands remain connected after replication
  • Semi-conservative model: Two parental strands each make a copy of itself, daughter molecules would each have one parental and one new strand after one round of replication
  • Dispersive model: Material in the two parental strands is dispersed randomly between the two daughter molecules, daughter molecules contain a random mix of parental and new DNA
• Meselson and Stahl experiment in 1954:

  • Used E. coli and heavy isotopes
  • Three-step process: cultured E. coli with heavy isotope N15, transferred bacteria to a medium with light isotope N14, centrifuged DNA and analyzed after each replication
  • Found that parental strands follow the semi-conservative model
• Steps in DNA replication:

  • Step 1: DNA replication begins at origins of replication, DNA opens to form replication fork
  • Step 2: Helicase unwinds DNA strands at each replication fork, single-stranded binding proteins (SSBPs) keep DNA open, topoisomerase prevents strain and damage above replication fork
  • Step 3: Primase adds short segments of RNA called primers to parental strand
  • Step 4: DNA polymerase 3 (DNA P3) attaches to each primer, moves in the 3 to 5 direction, synthesizes in the 5 to 3 direction
• DNA polymerase moves in one direction on the parental strand
• Nucleotides that DNA polymerase lays down are in the opposite direction, creating a new DNA strand that is anti-parallel
• DNA polymerase 3 following helicase is the leading strand and requires one primer
• DNA polymerase 3 on the parental strand moving away from helicase is the lagging strand and requires many primers
• Leading strand is synthesized in one continuous segment
• Lagging strand is synthesized in chunks known as Okazaki fragments
• After DNA polymerase 3 forms an Okazaki fragment, DNA polymerase 1 replaces RNA nucleotides with DNA nucleotides
• DNA ligase joins the Okazaki fragments forming a continuous DNA strand
• Telomeres protect genes on DNA from shortening on the lagging strand
• Telomeres are repeating units of short nucleotide sequences that do not code for genes and help postpone erosion
• Telomerase adds telomeres to DNA and is linked with aging
• DNA polymerase proofreads the bases added to the new DNA strand
• Mismatch repair removes and replaces incorrectly paired nucleotides
• Nuclease removes segments of damaged DNA if needed
• DNA polymerase and ligase can replace removed segments
• Proteins are polypeptides made up of amino acids linked by peptide bonds
• Gene expression is the process by which DNA directs the synthesis of proteins
• Gene expression includes two stages: transcription and translation
• Transcription is the synthesis of RNA using information from DNA
• Transcription occurs in the nucleus and allows the message of DNA to be transcribed
• Messenger RNA (mRNA) is synthesized during transcription using a DNA template
• mRNA carries information from the nucleus to the ribosomes in the cytoplasm
• Transfer RNA (tRNA) molecules carry specific amino acids and can attach to mRNA via an anticodon
• Ribosomal RNA (rRNA) helps to form ribosomes and link amino acids together
• DNA contains a sequence of nucleotides that code for proteins in groups of three called the triplet code