replication Flashcards
Process of duplication of DNA
replication
*requires RNA
Process of formation of RNA on a DNA template
transcription
*base sequence of DNA is reflected in the base sequence of RNA
[When RNA is formed from DNA during transcription, the order of bases (the genetic “letters”) in the DNA strand determines the order of bases in the new RNA molecule. It’s like the DNA is a template or a master copy, and the RNA is made to match it.]
Process of protein synthesis
translation
*aa sequence of the protein reflects the sequence of bases in the gene that codes for that protein
what is the flow of genetic information in nearly all organisms
DNA > RNA > protein
the genetic material is RNA and not DNA
retroviruses
*major exception
[retroviruses work backwards compared to the usual flow of genetic information (DNA to RNA to protein). They start with RNA, convert it to DNA, and then use the host cell’s machinery to replicate and spread. This unique feature makes retroviruses, such as HIV, particularly challenging to treat and study.]
it is the ability to turn their RNA into DNA once they infect a cell. This is the opposite of the usual process (where DNA is used to make RNA)
reverse transcription process
it is the key tool retroviruses use to perform this RNA-to-DNA conversion. It’s an enzyme that can read RNA and create a matching DNA strand.
reverse transcriptase enzyme
why does the retroviruses have to convert from RNA to DNA
to undergo self replication, hence the additional step
* for retroviruses only
what are the types of dna that it can exist in different forms in nature
Single-stranded or double-stranded
Linear (with two ends) or circular (forming a loop)
*because DNA comes in these different forms, there isn’t a one-size-fits-all explanation for how all DNA replicates. Each type might have some unique aspects to its replication process
it is common in a circular double-stranded DNA replicates
common in bacteria and some viruses
this is where most of the replication process were first investigated
in prokaryotes, particularly E. coli (a common gut bacterium)
why is it the single strands are vulnerable to damage when it is “unzipped” for replication
cell must protect these exposed strands
exposed to the cellular environment which makes it susceptible to various forms of damage or modification
exposed bases can more easily react with various molecules in the cell, potentially leading to chemical modifications
what unwound DNA needs protection from when it is “unzipped” for replication
nucleases
- these are enzymes that can break down single stranded DNA
free radicals
inappropriate base-pairings
how do you guard against errors in replication by ensuring that the correct base is added to the growing polynucleotide chain
DNA polymerase
- the main enzyme responsible for replication, has a built-in “proofreading” ability.
*
If an incorrect base is added, it’s usually detected immediately.
When an error is found, the incorrect base is removed and replaced with the correct one.
it is a DNA replication involves separation of the two original strands and production of two new
daughter strands using the original strands as templates
semiconservative replication
*each daughter strands contain
-1template strands (from DNA)
-1newly synthesizes strand
what bacteria is use where it was grown in an environment where the only nitrogen they could use came from a special form of nitrogen
E.coli
15N
- a heavier version of nitrogen
what method is used to track and study the bacteria’s nitrogen-containing molecules
density-gradient centrifugation
- this technique separates molecules based on their density. Since the 15N-labeled molecules are heavier than regular ones, they sink more in the centrifuge, making it easier to tell them apart from other molecules.
this the specific point where the DNA helix unwinds
origin of replication
true or false:
dna replication is bidirectional in most organisms
true
how many replication forks are there a each origin of replication
2
Points at which new polynucleotide chains are formed
replication forks
bidirectional growth of polynucleotide
means moving in two directions at the same time
In DNA replication, this means that the copying of DNA starts at one point and happens in both directions, going both left and right from that starting point. This helps the process go faster.
how many origins, replication fork and eye does prokaryotic and eukaryotic have
eukaryotic:
many origins
multiple replication fork
example - 3 origin, 6RF or 4 origin, 8 RF
eye depends on how many origin
prokaryotic:
1 origins
2 replication fork
1 eye
what direction does dna polymerase move towards
from 5’ to 3’
3’ has OH group which acts as the Nu = helps like the nucleotide tgt
when Nu is added
removed - pyrophosphate
formed - phosphodiester bond
Synthesized continuously from its 5′ end to its 3′ end at the replication fork on the exposed 3′ to 5′ template strand
leading strand
Synthesized semidiscontinuously in small fragments, or Okazaki fragments
lagging strand
*fragments are linked together by the enzyme DNA ligase
Enzymes that form DNA from deoxyribonucleotides on a DNA template
DNA Polymerases
in E. coli, how many DNA polymerases are there and how many have been extensively studied
5 DNA polymerase
3 have been extensively studied
(index card)
Number of nucleotides incorporated before the dissociation of enzyme from the template
processivity
* A high processivity means the enzyme can add a lot of nucleotides without stopping, which is helpful for efficiently building long DNA strands.
This refers to how fast a DNA polymerase can add nucleotides to the growing DNA strand.
Speed of synthetic Reaction (Turnover Number)
what is the turnover number and the processivity in pol I and II
turnover no:
II - 30
I - 600
III - 1200
processivity:
I - 200
II - 1500
III - more than 500,000
they are part of proofreading and repair functions
and what are the 2 functions
exonuclease activities
- function of some DNA polymerases that allows them to remove nucleotides from the ends of a DNA strand.
1) proofreading
Removing incorrect nucleotides during DNA replication
2) repair
Removing incorrect nucleotides from DNA and replacing them with correct ones
true or false:
DNA polymerase cannot catalyze de novo synthesis
false - cannot
de novo synthesis means that the process of creating something completely new from scratch, without using any existing templates or building blocks
what is a primer
a short piece of RNA
that acts as a starting point for DNA synthesis
attaches to the existing DNA strand (the template) through hydrogen bonds.
how does a primer work
primer has a free end with a 3′-hydroxyl group
DNA polymerase can only add new DNA nucleotides to this free end of the primer
new DNA strand begins to grow from there
it is the enzyme that makes primer
primase
what are the requirements for DNA polymerase reactions
Deoxyribonucleoside Triphosphates (dNTPs):
dTTP (for thymine)
dATP (for adenine)
dGTP (for guanine)
dCTP (for cytosine)
Ribonucleoside triphosphates (rNTPs):
ATP
UTP
GTP
CTP
*TTP is not applicable for RNA
Magnesium Ions (Mg²⁺): co factor
DNA Template
Ribonucleoside Triphosphates (rNTPs)
list all the replisomes that carries out dna replication
Complex of DNA polymerase
the RNA primer
primase
helicase
this is an enzyme that classified as a Class II topoisomerase. what are the functions
DNA gyrase
functions:
1. fights +ve supercoils (tightly twisted)
= helps relieve the twisting
- places -ve supercoils ahead of the replication fork
= helps keep the DNA unwound and ready for copying - energy required for the process is supplied by hydrolysis of ATP
-process of adding negative supercoils requires energy
- break down ATP to ADP
replisome vs DNA gyrase
replisome:
carries out the actual copying of DNA
DNA gyrase:
manages the twisting of the DNA by introducing negative supercoils, using energy from ATP to do so.
make transient single - stranded breaks in the DNA double helix and reseal the strands
topoisomerase I (Topo I)
make double-stranded breaks in the DNA double helix and reseal the strands using ATP
topoisomerase II (Topo II)
list all the primosome
RNA primer
primase
helicase
primosome vs replisome
primosome:
is a part of the replisome
focuses on getting DNA replication started by making the RNA primer
replisome:
responsible for the entire DNA replication process
briefly explain the synthesis of new strands using DNA polymerase III
it begins building new DNA strands by adding nucleotides - attaches the first nucleotide to the 3′-hydroxyl end of the RNA primer
in the 5′ to 3′ direction
- polymerase adds nucleotides to the 3′ end of the growing strand
happens on leading strand (continuously) and lagging strand (Okazaki fragments)
As the replication fork moves along, the RNA primer that started the process is no longer needed - hence, dna polymerase I removes the RNA primer and replace w DNA nucleotides
DNA ligase finishes the process
connecting any gaps between the newly made DNA pieces (especially in the lagging strand) = one continuous DNA strand
its is part of the DNA polymerase III complex, which is responsible for copying DNA during replication
clamp loader
* to open up a special protein called the sliding clamp and place it around the DNA strand.
sliding clamp:
hold DNA polymerase in place on the DNA, so it can move smoothly and efficiently while building the new DNA strand
clamp loader comes from a pentametric enzyme that is a member of a family of ATPases called
AAA+ superfamily
- powered by ATP
[clamp loader helps attach the sliding clamp to the DNA so that DNA polymerase III can stay attached and efficiently copy the DNA, using energy from ATP to get the job done]
Errors in replication that occur spontaneously only once in every 109–1010 base pairs and can be lethal to organisms
Mutations
Removal of incorrect nucleotides immediately after they are added to the growing DNA during replication
proofreading
what are the 2 main parts that Pol I can be cleaved into
klenow fragment:
1. Polymerase Activity
- helps build new DNA by adding nucleotides.
2. Proofreading Activity
- checks for mistakes and fixes them using its 3′ exonuclease activity, which can remove an incorrect nucleotide
other fragments:
5′ to 3′ Repair Activity
- remove damaged or unnecessary pieces of DNA (like the RNA primer) and replace them with the correct nucleotides
process where DNA Polymerase I fixes gaps in the DNA by removing damaged parts or the RNA primer and filling them in with the correct DNA.
nick translation
fixes gaps (called “nicks”)
function of Nick Translation
Cutting (Removal):
DNA Polymerase I uses its 5′ to 3′ exonuclease activity to cut out the RNA primer or any damaged parts of the DNA strand as it moves along.
Patching (Filling In):
After removing the incorrect or unnecessary parts, DNA Polymerase I uses its polymerase activity to fill in the gap by adding the correct DNA nucleotides.
Agents that bring about a mutation
Mutagens
what are the types of mutagens
mutagens - that can cause change
* Ultraviolet light, which creates pyrimidine dimers
* Ionizing radiation
* Various chemical agents or free radicals
what are the different type of DNA mechanism
Mismatch repair
Base-excision repair
Nucleotide-excision repair
Nonhomologous DNA end-joining (NHEJ)
*these mechanisms help keep the DNA intact by fixing mistakes and damage using different strategies based on the type of damage.
fixes mistakes that happen when the wrong bases are paired together during DNA replication.
mismatch repair
what is the challenge in mismatch repair
is figuring out which strand has the mistake. Right after DNA replication, there’s a small window of time when the repair system can recognize the new strand as the one with the mistake (since it’s the newly copied one).
Base-Excision Repair vs Nucleotide-Excision Repair
BER targets and removes single damaged bases, while NER removes a larger section of DNA that contains bulky damage.
In BER, a specific base is removed first, while in NER, an entire section of the DNA strand containing the damage is excised.
Natural process in which genetic information is rearranged to form new associations
genetic recombination
Involves a reaction between homologous sequences
Homologous recombination:
Involves combination of different nucleotide sequences
Nonhomologous recombination:
Occurs in specific zones of the chromosome. Specific areas on a chromosome where mutations are more likely to occur.
hot spots
