DNA Packaging

Question 1

explain bacteria genome’s supercoiling

Answer 1

• The circular DNA of bacteria genome is supercoiled as a result of the addition of turns being introduced into the DNA double helix
• Prokaryote DNA supercoiling is accomplished by the action of 2 enzymes:
  
  • DNA gyrase
  • DNA topoisomerase I

Question 2

explain the function of supercoiling

Answer 2

• Supercoiling compacts the DNA so that it can be packaged into the cell
• Negative supercoiling promotes strand replication (e.g. DNA replication)
• Positive supercoiling is introduced during DNA replication and transcription and must be corrected. Positive supercoiling makes DNA strand separation more difficult

Question 3

explain the function of ciprofloxacin

Answer 3

It inhibits DNA gyrase in bacteria

Question 4

explain the function of topoisomerase I

Answer 4

• It transiently forms a single covalent bond with the DNA and breaking a phoshodiester bond; this allows free rotation of the DNA around the covalent backbone bonds

Question 5

explain the function of topoisomerase II (DNA gyrase)

Answer 5

• This enzyme is activated when 2 double helices cross over each other
• Process:
  
  • 1. DNA gyrase makes a covalent linkage to both strands of one DNA helix and making a double stranded break
  • The second DNA helix is passed through the break
  • The break is resealed and the enzyme dissociates
• Requires ATP hydrolysis

Question 6

explain the organization of DNA

Answer 6

• Chromosomes are composed of chromatin
• Chromatin is the complex of DNA and chromosomal proteins
• The most important chromosomal proteins are called histones

Question 7

describe the nucleosome structure

Answer 7

• Various histone proteins bind with each other along with DNA to form a structure called the nucleosome. The nucleosome core is composed of 2 molecules each of H2A, H2B, H3 and H4 histone proteins

Question 8

explain the core octamer of the nucleosome

Answer 8

• Each nucleosome contains a core of histones around which DNA is wrapped, called the core particle of octamer
• Made up of octamer of core histones
  
  • 2 molecules of H2A, H2B, H3 and H4
• Positively charged amino acids of histone interact with negatively charged phosphate groups in DNA
• As the DNA is wrapped around the core particle it develops a single negative supercoil

Question 9

describe the structure of the nucleosome

Answer 9

• Includes the core octamer, linker DNA and histone H1
  
  • Nucleosomes and the linker DNA form the 1st level of packaging = chromatin (beads on a string)
    
    • 10 nm in diameter

Question 10

explain DNA methylation

Answer 10

• DNA methylation involves the modification of the DNA base cytosine to 5-methyl-cyotsine via DNA methyltransferase enzyme

Question 11

explain the function of DNA methylation

Answer 11

• DNA methylation tags a region of the genome for histone modification that causes the formation of heterochromatin (HC).
• Process:
  
  • 1. Dnmt3 methylates unmethylated DNA
  • MeCP2 recruits HDAC1
  • HDAC1 deacetylates histones
  • HC is formed
• Decitabine: causes DNA hypomethylation and increased transcription

Question 12

describe histone acetylation

Answer 12

• Acetylation removes the positive charges, which allows it to be less attached to negatively charged histones
• Increases transcription