7.1 DNA structure and replication Flashcards
What proteins is eukaryotic DNA vs. prokaryotic DNA associated with?
-Eukaryotic DNA is always associated with basic (alkaline) and positively charged proteins called histones.
-This is in contrast to prokaryotic DNA, which lacks histones, and is therefore often referred to as ‘naked’ DNA.
Describe the structure of a nucleosome
-A nucleosome consists of a length of DNA of about 150 base pairs, wrapped around a core of eight histones (which are actually four pairs of four different histones) and a special histone named H1.
-DNA is acidic and negatively charged, so the bonding with the histones neutralises the DNA.
Diagram of a nucleosome
Why are nucleosomes linked and what is this called?
-The nucleosomes are linked because the DNA strand from one nucleosome flows directly into the next nucleosome.
-This section of DNA is called a DNA linker.
-The overall appearance of DNA in this form has been called a string of ‘beads’.
Diagram showing the nucleosomes, the DNA fibre and a chromosome
Why is a certain amount of packaging (folding, coiling, and re-coiling) required to fit the genetic material into the nucleus?
Because some eukaryotes have large genomes.
How is packaging done to fit the genetic material into a eukaryotic nucleus?
-Nucleosomes help to supercoil the DNA while still ensuring appropriate access to it.
-Access to DNA occurs when the coils unwind and histones are moved out of the way so that DNA can be copied or transcribed.
-Nucleosomes can be considered to be the repeat units of eukaryotic chromatin, which is further coiled to form chromosomes.
What does a eukaryotic chromosome consist of?
A single linear molecule of double-stranded DNA plus proteins.
What is structure X in the diagram of a nucleosome?
Histone
What is the function of the nucleosome?
To supercoil DNA
What type of cells have DNA associated with histones?
Eukaryotic
What does DNA replication rely on?
Base pairing
Give an overview of how Watson and Crick eventually worked out base pairing (reword?)
-It took Watson and Crick a long time to work out base pairing.
-They were trying to assemble a compact molecule of DNA because the X-ray diffraction work of Franklin suggested this type of model.
-The DNA molecule also needed to be stable because of its function as genetic material.
What properties does base-pairing allow the double helix to have?
-The hydrogen bonding between the purine and the pyrimidines.
-Two hydrogen bonds occur between adenine (A) and thymine (T), and three hydrogen bonds occur between guanine (G) and cytosine (C).
-The slightly positive charge on T and a slightly negative charge on A allow the two bases to bond together during complementary base pairing.
How was a mechanism for DNA replication presented? (reword?)
-Once it became clear that DNA forms a double helix with antiparallel strands bonded together by complementary base pairing, a mechanism for DNA replication also presented itself.
-If the double helix of a single DNA molecule was separated, each strand could be used to create the matching new strand through complementary base pairing, resulting in two new identical molecules.
-If, on one strand, the bases T and C occurred successively, automatically on the opposite strand A and G would be present, because A can only pair with T and C can only pair with G.
Diagram of double-stranded DNA showing antiparallel strands and base pairing
What are purines and pyrimidines and how do they bond in base pairing?
-Purines are: Guanine and Adenine (they contain two rings in their structure).
-Pyrimidines are: Thymine and Cytosine (they contain one ring in their structure).
-In DNA base pairing, a pyrimidine is always bonded to purine.
How does DNA replication progress?
-In a semi-conservative way.
-However, the details of DNA replication differ between eukaryotes and prokaryotes.
How does DNA replication differ between eukaryotes and prokaryotes?
-The main difference is that in eukaryotes, replication can be initiated at various points along the DNA molecule, while it can be started at only one position on prokaryotic DNA.
-This ensures more efficient DNA replication in eukaryotes.
-You are expected to study the prokaryotic system only.
Diagram showing DNA replication in prokaryotes versus DNA replication in eukaryotes
Describe the role of enzymes in DNA replication
-DNA replication is carried out by a complex system of enzymes.
-The most important ones are helicase, DNA gyrase, DNA primase, DNA ligase, as well as DNA polymerase I and III.
-The role played by each one of these enzymes in DNA replication should be known.
More info needed on rates of replication and size of human genome?
-The rate of replication is approximately 100 nucleotides per second in eukaryotes while it can be as high as 1000 nucleotides per second for prokaryotes.
-The human genome has around 3 billion base pairs per haploid set of chromosomes, so 6 billion base pairs have to be replicated during the S phase of the cell cycle.
How does efficiency change when a DNA molecule is replicated?
When a DNA molecule is replicated, efficiency is improved if it progresses in two directions.
DNA replication requires two ___
Replication forks
Diagram of semi-conservative DNA replication
How do helicase, SSBPs, and gyrase work together?
-First of all, helicase binds to the origin of replication and breaks hydrogen bonds between base pairs to unwind the DNA double helix.
-Single-strand binding proteins then bind to the single strands formed to keep them apart to allow time for the DNA sequence to be copied.
-The two separated strands act as templates for the replication process.
-As helicase moves along the DNA molecule, it causes supercoiling and tension on the region ahead.
-This is relieved by the enzyme DNA gyrase, which moves in advance of helicase.
What happens to free nucleoside triphosphates when they bind to their template (during DNA replication)?
They lose their two extra phosphate groups to generate energy, which is used to add the nucleotide to the growing polynucleotide chain.
What is the leading strand?
The strand of DNA that is being replicated continuously in the 5’ to 3’ direction by continuous polymerisation at the 3’ growing tip.
What is the lagging strang?
The strand of DNA that is replicated discontinuously in small fragments in the 5’ to 3’ direction away from the replication fork.
What happens on the leading strand during DNA replication?
-DNA polymerase III, the enzyme that adds DNA nucleotides to the strands, can only add a nucleotide to the 3’ OH group of the deoxyribose.
-So, on the leading strand, the DNA polymerase follows the helicase, separating the strands and adding the DNA nucleotides.
-Because no DNA polymerase enzyme can initiate a new DNA on its own, an RNA primer is needed once for this leading strand
What happens on the lagging strand during DNA replication?
-Because no DNA polymerase enzyme can initiate a new DNA on its own, an RNA primer is needed once for this leading strand.
-However, this is not possible on the lagging strand, because the last nucleotide ends with a 5’ phosphate group.
-So, on the lagging strand, a DNA primase first makes short RNA primers (these primers are later removed by DNA polymerase I and substituted with a short DNA segment), which allow the DNA polymerase III to add DNA nucleotides to the 3’ OH of the RNA primer.
-Many such primers are made as a scaffold for the DNA polymerase III.
-It synthesizes short DNA fragments called Okazaki fragments.
-The result is two new strands, both based on the template of the old DNA molecule.
What are Okazaki fragments?
Short DNA fragments, which are joined together by DNA ligase to form a complete DNA strand.
Why does the leading strand require one primer?
Since DNA polymerase III needs a 3’ OH group to start the polymerisation process, even the leading strand requires one primer.
What are RNA primers?
Short RNA chains of about 10 bases used as a starting point for DNA replication by DNA polymerase III. They are synthesised by DNA primases.
In what direction does DNA replication progress and why?
DNA replication always progresses from 5’ to 3’, because DNA polymerase III ( three ) can only add nucleotides to the 3’ OH group of the deoxyribose.
Cytosine makes up 42% of the nucleotides in a sample of DNA from an organism.
Approximately what percentage of the nucleotides in this sample will be thymine?
8%
If there is 42% of C then there must be 42% of G as well since G pairs with C. That leaves 16% for A and T together. So, T is 16÷2=8%.
What is meant by the description ‘antiparallel’ regarding the strands that make up DNA?
The 5’ to 3’ direction of one strand runs counter to the 3’ to 5’ direction of the other strand.
What components does an Okazaki fragment have?
RNA primer, DNA fragment
A biochemist isolated and purified molecules needed for DNA replication.
When she added some DNA, replication occurred but the DNA molecules were defective.
Each consisted of a normal DNA strand paired with numerous segments of DNA a few hundred nucleotides long.
What had she probably left out of the mixture?
-DNA ligase
-The scientist had DNA molecules that consisted of unlinked Okazaki fragments in one half of the molecule.
-DNA ligase is necessary to join the Okazaki fragments together on the lagging strand.
What is the reason that the leading and lagging strands of DNA are synthesised differently?
DNA polymerase III can only join new nucleotides to the 3’ end of the growing strand.
Which enzyme unwinds the DNA double helix during replication?
Helicase
Explain how every base in a DNA molecule is part of a gene
Non-coding sequences are found everywhere in eukaryotic genomes, and some genomes have more than others.
How much of the human genome does non-coding DNA account for?
More than 98%
Image of telomeres located at each end of the chromosome (false color image of a chromosome)
Diagram of the variation of tandem repeats in three different individuals
Diagram showing DNA profiling of two individuals and how they differ in the number of tandem repeats and how that is visualised (shown) on a gel (reword?)
Describe the tandem repeats for each individual in this diagram of a DNA profile
-In Individual #1, you can see that for allele A6 there are five repeats of the GC tandem (10 base pairs), but for allele A2, there are only two repeats, so that allele is shorter (4 base pairs).
-When these two alleles are separated by electrophoresis, the larger A6 segment is separated from the smaller A2 segment.
-These differences can be clearly seen on the gel.
-The pattern on the gel of each individual’s alleles at each locus forms the DNA fingerprint.
The DNA profiles that follow represent four different individuals.
Who is the child of whom?
-C is the child of A and B.
-All the DNA fragments for C can be traced to either A or B.
Diagram of two different nucleotides- deoxyadeosine and dideoxyadenosine
Diagram showing the basics of the dideoxy chain terminating sequencing method
Image of X-ray diffraction of DNA
It is from a DNA sample that came from Watson and Crick’s labs in Cambridge University, UK.
The radiogram shows the distance between the base pairs, as well as the turns of the helix (3.4 nm).
Franklin was able to measure other features of the DNA molecule, including the distance between the repeats.
Image of bacteriophages (virus) injecting DNA into a bacterial cell
Diagram of the Hershey-Chase experiment
The diagram displays the setup of part of the Hershey–Chase experiment. Once the viruses have infected the bacterium, and they are destroyed and separated at the end of the experiment, where would you expect to find the radioactivity?
In the bacterium
