Chapter 7 - Genomes and Chromosomes
● Supercoiling/Negative supercoiling
- Form: circular DNA can be broken and twisted to form supercoils
- Negative supercoil – loss of a twist in the DNA helix which causes DNA to
compensate by twisting around itself
- Type 1 topoisomerase: one negative supercoil is removed
- Type 2 topoisomerase: add one negative supercoil
- gyrB: holds onto Dna
- gyrA: breaks dna strand, open it, and reseals it adding negative supercoil.
- Topoisomerase 4: unlinks the two chromosomes after replication
● DNA replication
1. DnaA binds to oriC
2. DnaA causes strand separation by folding the DNA
3. DnaC helps attach DnaB (helicase) to the DNA strands
4. Helicase unwind DNA in both directions
5. DNA primase adds RNA primers for the leading strand
6. Clamp loaders bring DNA Polymerase III to the leading strand. Helicase
unwinds more DNA and lagging strand primers are made
7. Leading strand elongation begins
➔ Elongation Part 1
1. Single stranded DNA binding proteins protect ssDNA
2. Lagging strand is elongated
3. Every 1000 bases a new lagging strand primer is made
4. Lagging strand polymerase comes off after bumping into the previous
fragment
5. New clamp attaches to new RNA primer on lagging strand
6. Next Okazaki fragment is made
● Plasmid replication
- Rolling circle model:
- RepA cuts open one strand of the plasmid at the ori
- DNA polymerase III replicates the cut strand while old strand comes off
- RepA cuts apart the completed and single stranded plasmids
- DNA polymerase finished the single-stranded plasmid
● Eukaryotic vs prokaryotic chromosomes
- Eukaryotic chromosomes characteristics:
- Packaged around histone proteins
- Contains introns
- Contains many noncoding regions
- Archaeal chromosome characteristics:
, - Contain a single circular chromosome
- Bidirectional replication (two forks)
- Multiple origins of replication
Chapter 8
● Sigma factors and promoters
- Promoters: regions of DNA upstream (before) the transcription start site
where holoenzyme binds
- Location of most promoters: 35 and 10 nucleotides before the beginning of
the transcription start site
- Sigma: RpoD (sigma 70) – the most common sigma factor in E. coli and
activates most genes
- Sigma factor binding to promoter happens be binding to the -35 and -10
regions on the promoter
● What is a gene
- Untranslated leader – Beginning of mRNA molecule that contains the
ribosome binding site
- Transcription terminator – regions at the end of the DNA sequence/gene to
stop transcription
● Transcription initiation and termination
- Initiation
1. Holoenzyme scans the DNA molecule and finds promoter
2. Holoenzyme binds to the promoter
3. Ribonucleotides arrive and transcription begins – sigma factor comes off
- Termination:
1. At the stop codon, Release factor 1 or 2 binds to A site
2. Created protein is released
3. RF3 pushes out RF1 or 2
4. RRF and EF-G-GTP uses GTP energy to split 50S and 30S subunits
5. IF3 binds to 30S subunit
● tRNA function
- carries a specific amino acid to the ribosome during protein synthesis
● Translation./Protein Synthesis steps
➔ Synthesis Initiation
1. Initiation factor 3 splits 30S and 50S subunit
2. 30S binds to the ribosome binding site (Shine-dalgarno) sequence on
mRNA and IF1 binds to A site
3. IF2 brings Met-tRNA to the P site
4. IF 1, 2, and 3 leave
5. 50S subunit binds
● Supercoiling/Negative supercoiling
- Form: circular DNA can be broken and twisted to form supercoils
- Negative supercoil – loss of a twist in the DNA helix which causes DNA to
compensate by twisting around itself
- Type 1 topoisomerase: one negative supercoil is removed
- Type 2 topoisomerase: add one negative supercoil
- gyrB: holds onto Dna
- gyrA: breaks dna strand, open it, and reseals it adding negative supercoil.
- Topoisomerase 4: unlinks the two chromosomes after replication
● DNA replication
1. DnaA binds to oriC
2. DnaA causes strand separation by folding the DNA
3. DnaC helps attach DnaB (helicase) to the DNA strands
4. Helicase unwind DNA in both directions
5. DNA primase adds RNA primers for the leading strand
6. Clamp loaders bring DNA Polymerase III to the leading strand. Helicase
unwinds more DNA and lagging strand primers are made
7. Leading strand elongation begins
➔ Elongation Part 1
1. Single stranded DNA binding proteins protect ssDNA
2. Lagging strand is elongated
3. Every 1000 bases a new lagging strand primer is made
4. Lagging strand polymerase comes off after bumping into the previous
fragment
5. New clamp attaches to new RNA primer on lagging strand
6. Next Okazaki fragment is made
● Plasmid replication
- Rolling circle model:
- RepA cuts open one strand of the plasmid at the ori
- DNA polymerase III replicates the cut strand while old strand comes off
- RepA cuts apart the completed and single stranded plasmids
- DNA polymerase finished the single-stranded plasmid
● Eukaryotic vs prokaryotic chromosomes
- Eukaryotic chromosomes characteristics:
- Packaged around histone proteins
- Contains introns
- Contains many noncoding regions
- Archaeal chromosome characteristics:
, - Contain a single circular chromosome
- Bidirectional replication (two forks)
- Multiple origins of replication
Chapter 8
● Sigma factors and promoters
- Promoters: regions of DNA upstream (before) the transcription start site
where holoenzyme binds
- Location of most promoters: 35 and 10 nucleotides before the beginning of
the transcription start site
- Sigma: RpoD (sigma 70) – the most common sigma factor in E. coli and
activates most genes
- Sigma factor binding to promoter happens be binding to the -35 and -10
regions on the promoter
● What is a gene
- Untranslated leader – Beginning of mRNA molecule that contains the
ribosome binding site
- Transcription terminator – regions at the end of the DNA sequence/gene to
stop transcription
● Transcription initiation and termination
- Initiation
1. Holoenzyme scans the DNA molecule and finds promoter
2. Holoenzyme binds to the promoter
3. Ribonucleotides arrive and transcription begins – sigma factor comes off
- Termination:
1. At the stop codon, Release factor 1 or 2 binds to A site
2. Created protein is released
3. RF3 pushes out RF1 or 2
4. RRF and EF-G-GTP uses GTP energy to split 50S and 30S subunits
5. IF3 binds to 30S subunit
● tRNA function
- carries a specific amino acid to the ribosome during protein synthesis
● Translation./Protein Synthesis steps
➔ Synthesis Initiation
1. Initiation factor 3 splits 30S and 50S subunit
2. 30S binds to the ribosome binding site (Shine-dalgarno) sequence on
mRNA and IF1 binds to A site
3. IF2 brings Met-tRNA to the P site
4. IF 1, 2, and 3 leave
5. 50S subunit binds
