Protein Synthesis and DNA Replication - Genes Flashcards
What is DNA replication and how often foes it occur in a cell cycle?
Producing two identical DNA molecules from one DNA molecule - it must happen only once per cell cycle
What is the enzyme that synthesizes DNA and in what direction does it do this?
DNA polymerase III (Pol III) - 5’ to 3’ direction
What does highly processive mean when referring to Pol III?
Once it starts the enzyme doesn’t dissociate until it is terminated
What is an Okazaki fragment and why are they formed?
Pol III has no issue continuously replicating the leading strand as it is in the 5’ to 3’ direction. However the lagging strand is in the 3’ to 5’ direction so a series of ~1500 base fragments are replicated in the 5’ to 3’ direction before being joint togther
What does semi-discontinuous DNA replication refer to?
The leading strand is replicated continuously but the lagging strand is replicated in Okazaki fragments discontinuously
What is a primer?
A piece of RNA that is used as a starting point as Pol III can not begin synthesis alone. For the leading strand only one is require but many are required for the lagging strand
What is the enzyme that is responsible for making primers?
DNA primase
What are the functions of DNA polymerase 1 and DNA ligase?
DNA polymerase 1 - replaces RNA primers with DNA
DNA ligase - joins the newly replicated DNA on the lagging strand together
What is the function of DNA helicase?
It unwinds duplex DNA to single stranded DNA
What does in mean if DNA is positively supercoiled?
It is overwound
What is a linking number and what is its equation?
The number of times one strand winds around the other
Lk = Lk0 = N/h (for relaxed DNA)
Where:
N = number of base pairs
h = 10.5 (helical repeat in solution)
What is the equation for specific linkage difference (σ)?
σ = ΔLk/Lk0 where ΔLk = Lk - Lk0
Does in vivo DNA have a higher or lower linking number than in vitro?
DNA in cells is negatively supercoiled (underwound) and so it has a lower linking number than synthesized DNA
What are DNA topoisomerases?
Enzymes that break closed circular DNA to resolve topological complexity (over- or underwound DNA) and can pass remote DNA through the break
How can the linking number of a closed circular DNA be changed without breaking the DNA?
It can’t !!!!
What is the initiation of DNA replication called?
oriC
What protein binds to DNA for replication to begin?
DnaA
In which direction does DNA replication occur?
It is bidirectional
What are the steps involved in DNA replication initiation?
1) Binding of DnaA to DnaA boxes at oriC
2) Unwinding of DNA double helix by a DNA helicase
3) DNA primase synthesizes RNA primers
4) Pol III extends RNA primers with DNA
What are the “ingredients” used in Sanger sequencing of DNA?
- single stranded DNA template (usually from a plasmid)
- primer
- DNA polymerase (Pol III)
- dATP, dCTP, dTTP, dGTP
- radioactive dATP
- one of ddATP, ddCTP, ddTTP, ddGTP (for termination)
Why does ddATP result in termination?
The missing the 3’ OH group results in termination and therefore if the template is T and requires an A there is a chance that A will come from ddATP and cause termination
How does polyacrylamide gel electrophoresis work?
Negative DNA moves due to an electric current with smaller strands of DNA moving more quickly towards the positive charge. They are read by moving up and seeing which base is next
In which direction do peptides grow?
From the N-terminus to the C-terminus
Does DNA replication or translation have a higher error frequency?
Translation has a higher error frequency as the consequence of one protein having an incorrect amino acid is far less than that of DNA having an error which will result in many proteins with errors for many generations
