What are the structural features of DNA?
- Double Helix
- Four Bases
- Hydrogen Bonding between bases
- A-T & G-C
- Antiparallel alignment
List and describe the methods of replication of DNA.
- Dispersive : Fragments of the original are dispersed throughout the new DNA strand.
- Conservative : New DNA strand contains none of the original DNA.
- Semi-Conservative : Each Helix of DNA has one strand from the original DNA.
Describe the direction of synthesis of bacterial daughter DNA.
It is bidirectional.
What is the origin of replication called in E. Coli?
oriC (Origin on Chromosomal Replication)
What are the three types of DNA sequences in the oriC?
- AT rich region
- DnaA box (5)
- GATC Methylation Site
What binds to the five DnaA boxes?
What takes place after DnaA proteins bind to the five DnaA boxes?
Stimulation occurs, which causes cooperative binding of additional 20-40 DnaA proteins to form a large complex.
What causes the AT-rich region to seperate in initiation of replication?
Tension of DNA being wrapped around DnaA protein complexes.
Following denaturation of AT-rich regions in bacterial DNA replication, what promotes helicase enzymes (DnaB helicase) to begin seperation within oriC?
- DnaA proteins
- DnaC proteins
Does the action of breaking Hydrogen Bonds by DnaB helicase in bacterial DNA replication require energy?
- It hydrolyzes ATP for energy.
What enzyme alleviates supercoiling in the strand as helicase splits apart DNA in bacterial DNA replication?
- DNA gyrase
What is the function of SSB proteins in bacterial DNA replication?
These bind to the strands of parental DNA and prevent them from reforming a double helix until complementary daughter strands can be formed.
What enzyme synthesizes the leading strand of DNA?
DNA polymerase 3
Describe the proces of DNA replication of the lagging strand in bacterial DNA replication.
- Primase synthesizes RNA primer so that Polymerase 3 can bind to the primer and begin.
- Polymerase 3 elongates the primer (producing an okazaki fragment). Then leaves strand.
- Polymerase 1 excises RNA primer and fills the gap.
- DNA ligase covalently links the okazaki fragments to form continuous strand.
What type of bond is formed between nucleotides by DNA polymerases in daughter strands of bacterial DNA replication?
What DNA polymerases are involved in normal DNA replication in bacterial DNA replication?
- DNA polymerase 1
- DNA polymerase 3
Describe the structure and function of DNA polymerase 1.
- Single subunit
- Fills in small regions where RNA primers were located.
Describe the sructure and function of DNA polymerase 3.
- 10 subunits
- Beta clamp : Wraps the polymerase around the strand to lock the enzyme onto the strand.
- Alpha : Synthesizes DNA.
- Theata and epsilon : Proofread nucleotides.
- It is responsible for most of DNA replication.
- A processive enzyme.
What is a holoenzyme?
A multi-subunit enzyme.
What are the two oddities of DNA polymerase in bacterial cells?
- Unable to initiate DNA synthesis by linking togehter two individual nucleases.
- Is not bidirectionally synthetic.
Where are TER sequences located on the bacterial chromsome?
Opposite of the oriC.
What binds to the TER sequences to stop the replication fork?
Termination utilization substance (TUS) proteins
What is the function of T1 and T2 TER sequences in the bacterial chromosome?
- T1 : allows advancement of clockwise moving forks, but prevents the advancement of counterclockwise moving forks.
- T2 : allows advancement of counterclockwise moving forks, but prevents the advancement of clockwise moving forks.
What links the two daughter strands creating two circular double stranded molecules in bacterial DNA replication?
What is a Cantenane?
Two interlocked circular DNA strands.
In E. Coli, what breaks cantenanes and rejoins the unlocked DNA?
What is a Primosome?
DNA helicase + Primase
What is a Replisome?
Primosome + two DNA polymerases holoenzymes
What is Dimeric DNA polymerase?
Two polymerase holoenzymes that move as a unit toward the replication fork.
What are the reasons that DNA replication exhibits a high degree of accuracy?
- Hydrogen bonds betwen correct nucleotide partners are more stable.
- Exonucleases remove mismatch bases at 3' end of a newly made strand.
- Subunit epsilon, alpha, and theata identify and remove mismatched nucleotides in the 3' to 5' direction.
What are the two different initiation mechanisms in bacterial cells?
- 1) Insufficient amount of DnaA proteins to bind to all DnaA boxes within the oriC, thus preventing premature replication.
- 2) Temporary lack of adenine methylation in the GATC sites within the oriC. Dam enzymes take several minutes to methylate the DNA for replication to start.
What does the term Hemimethylation refer to?
- Methylation on one strand.
- DNA must be full methylated in bacterial cells for replication to begin.
What replication enzymes do eukaryotes share with prokaryotes?
- Ligase (Topoisomerase)
How many Origins of replication do eukaryotes have?
Describe the direction of replication of DNA in eukaryotes.
- It is bidirectional from many origins of replication.
- Takes place in S phase.
- Forks eventually meet and complete the process.
What are ARS elements?
- Autonomously replicating sequences
- High AT content
- ATTAT (A or G) TTTA
- Equivilent to AT-rich area of bacteria
What is the ORC?
- Origin of recognition complex
- Six subunit protein complex that acts as the initiator of eukaryotic DNA replication when it binds to eukaryotic DNA replication when it binds to ARS during G1 phase.
How many different DNA polymerases are present in a eukaryotic cell?
Where can DNA polymerase enzymes be found in a eukaryotic cell?
What do bacteria and Eukaryotes use to remove RNA primers?
- Bacteria : DNA Polymerase 1
- Eukaryote : Flap endonuclease
When does the synthesis of new histone proteins take place?
Describe the despersion of histone proteins between daughter strands of DNA.
Each strand contains a random mixture of original and newly assembled octamers.
What is a Telomere?
Moderatly repetitive random array with a 3' overhang region (12-16 nucleotides)
Describe the composition of Telomerase.
- It is a Ribonucleoprotein.
- Contains both proteins and RNA (complementary to the telomeric repeats at the end of the chromosomes).
- Its RNA molecule is calle TERC (Telomere RNA Component)
- Its protein component is called TERT (Telomere Reverse Transcriptase)
Describe the function of Telomerase.
is a reverse transcriptase that synthesizes DNA from an RNA template.
What is polymerization?
Attachment of a series of six-nucleotide sequences (tandom repeats) to the end of a DNA strand.
Is Telomerase found in all eukaryotic cells?
- It is only found in few somatic cells.