Chapter 16: DNA Flashcards
Transformation
Regarding genes
A change in genotype and phenotype due to assimilation of foreign DNA
Discovered by Frederick Griffith in 1928
Bacteriophages
Viruses that infect bacteria
Phages for short
Hershey and Chase experiment
Designed an experiment to see which components of a phage known as T2 enters an E.coli during infection
Determined that DNA is the genetic material
Chargaff’s rules
- DNA base composition varies between species
- In each species the percentages fo A and T bases are roughlyl equal as well as those of C and G
Structure of DNA strand
Each DNA nucleotide monomer consists of three components:
- A sugar deoxiribose group
- A nitrogenous base (A, T, G, C)
- A phosphate group
Bases are stacked every 0.34 nm apart
Helix makes one full turn every 3.4 nm along its length; every 10 base pairs
Polynucleotide strand has built-in directionality from the 5’ end to the 3’ end
- 5’ terminal end has a lone phosphate group attached to the 5’ carbon
- 3’ terminal end has an -OH group attached to the 3’ carbon
Nucleotide base pairing
Nitrogenous bases are attached to the 1’ carbon
A purine- adenine and guanine, must pair with a pyrimidine- cytosine and thymine
Hydrogen bonds between base pairs hold the strand together
- 2 hydrogen bonds pair adenine to thymine
- 3 hydrogen bonds pair cytosine to guanine
Van der Waals interactions between stacked base pairs help hold the molecule together
Model for DNA replication
Semiconservative model- after replication the two daughter molecules will have one old parental strand and one new strand
Meselson and Stahl experiment
Experiments by Matthew Meselson and Franklin Stahl supported the semiconservative model
They labeled the nucleotides of the old strands with a heavy isotope of nitrogen and new nucleotides were labeled with a lighter isotope
- The first replication produced a band of hybrid DNA, eliminating the conservative model
- A second replication produced both light and hybrid DNA, eliminating the dispersive model and supporting the semiconservative model
Origins of replication
Site where DNA replication begins
Proteins that initiate replication recognize a specific sequence and attach to the DNA opening up a replication “bubble”
At the end of each replication bubble is a replication fork- a Y-shaped region where new DNA strands are elongating
Replication then proceeds in both directions until completion
Prokaryotes only have one origin while eukaryotic DNA can have hundreds or even a few thousand
Proteins that participate in unwinding
Helicases- enzymes that untwist the double helix at the replication forks separating the two parental strands
Single-strand binding proteins- bind to the unpaired DNA strands and keep them from re-pairing
Topoisomerase- enzyme that helps relieve the tighter twisting that occurs ahead of the replication fork due to the untwisting of the double helix; breaks, swivels and rejoins DNA strands
Preparation for replication
DNA polymerase cannot initiate the synthesis of polynucelotides; can only add to existing chain that is base paired to the template strand
Initial nucleotide chain is a short stretch of DNA generally 5 to 10 nucleotides long called a primer
Primer is synthesized by the enzyme primase
The new DNA strand will start from the 3’ end of the RNA primer
DNA polymerase
Enzymes that catalyze the sythesis of DNA by adding nucleotides to the 3’ end of a preexisting chain; new strand elongates in the 5’ → 3’ direction
Require a primer and a DNA template strand
In prokaryotes such as E. coli several DNA polymerases play a role however DNA polymerase III and I play the largest; add about 500 nucleotides per second
In eukaryotes it is more complicated with at least 11 DNA polymerases discovered thus far; add about 50 nucleotides per second in humans
Leading strand elongation
Along the leading strand DNA polymerase can sythesize a complementary strand continuously in the required 3’ → 5’ direction towards the replication fork as it progresses
Only one primer is required to sythesize the entire leading strand
Lagging strand elongation
Along the lagging strand DNA polymerase has to sythesize the complementary strand away from the replication fork to still go in the required 3’ → 5’ direction
The lagging strand is synthesized discontinuously as a series of segments called Okazaki fragments
Whereas only one primer is required for the leading strand, each Okazaki fragment requires a seperate primer
After DNA polymerase III forms an Okazki fragment, another DNA polymerase, DNA polymerase I replaces the RNA primers with DNA one at a time
DNA polymerase I cannot join the replacement DNA to the Okazaki fragment; another enzyme called DNA ligase is required to join the sugar phosphate backbones of all the Okazaki fragments into a continuous DNA strand
DNA replication complex
The various proteins that participate in DNA replication form a single large complex; a “DNA replication machine”
DNA is pulled through the complex which is anchored in the nuclear matrix
Proofreading and repairing DNA
Several mechanisms are responsible for proofreading and repairing DNA
- Many DNA polymerases proofread each nucleotide againstits template strand as soon as it is covalently bonded to the growing strand
- Mismatch repair- other enzymes remove and replace incorrectly paired nucleotides
- Nucleotide excision repair- a segment of damaged DNA is excised by a DNA-cutting enzyme called a nuclease; resulting gap is filled in by DNA polymerase and ligase
Replicating the ends of DNA molecules
The usual replication machinery provides no way to complete the 5′ ends because there is no preexisting 3’ end for DNA polymerase to add on to
In eukaryotes repeated rounds of replication produce ever shorter DNA molecules with uneven ends
Eukaryotic DNA thus has special nucleotide sequences on its ends called telomeres which consist of multiple repititions of one short nucleotide sequence that does not contain coding genes; TTAGGG in humans
Telomeres have two protective funcitons
- Prevent the staggered ends of daughter strands from activating the cell’s systems for monitoring DNA damage
- Acts as a kind of buffer zone that provides some protectiong against the shortening of and organism’s genes
An enzyme called telomerase catalyzes the lengthening of telomeres in germ cells to restore them to their original length
DNA packaging
During interphase in eukaryotes
DNA is precisely combined with proteins in a complex called chromatin
Proteins called histones are responsible for the primary level of DNA packing of interphase chromatin
- More than a fifth of a histone’s amino acids are positively charged lysine and arginine which bind tightly to the negatively charged DNA
- Five types: H1, H2a, H2b, H3, and H4
Unfolded DNA resembles “beads on a string” with each bead representing a nucleosome and the string representing linker DNA
- A nucleosome is the basic unit of DNA packing; consist of DNA wound twice around a protein core of eight histones
- Amino end of each histone extends owtwards from the nucleosome and is involved in gene regulation and gene expression
Chromatin
In eukaryotes
Complex of DNA and a large amount of protein percisely combined in eukaryotic cells
Two main types:
- Euchromatin less compacted 10 nm strand that is more dispersed during interphase; genes are accessible to transcription proteins and can be expressed
- Heterochromatin a more compacted, denser-appearing 10 nm strand whose genes are not generally expressed; e.g. centromeres and telomeres
Both joined by linker DNA
Chromosomal packing
During the mitotic phase in eukaryotes
Two proteins play a role in DNA condensing during prophase and prometaphas of mitosis
Condensin II (red) binds to the 10 nm chromatin fiber in prophase and forms loops that get larger and larger; form a central scaffold from which the loops extend
Condensin I (green) binds to the chromatin outside of the central scaffold during prometaphase making smaller loops out of the larger loops; process continues with more and more loops extending outward making the chromosome progressively denser and wider
