Exam review
leading strand
the new DNA polymer at the opening of the replication fork (DNA is building from the 3' end
lagging strand
the new DNA polymer that is opposite the opening of the replication fork (building from 3' end)
What is DNA replication?
DNA replication occurs before cell division and is required for reproduction, growth, and tissue replacement. The Original DNA strands serves as a template for the creation of a new strand. DNA polymerase 3 builds the new strand by reading the template and adding the complementary DNA nucleotide.
Initiation (DNA replication)
begins at a specific nucleotide sequence called the origin of replication (creates replication bubbles all over the DNA) . Origin recognition complexed bind to the DNA to begin unwinding it. DNA helicase unwinds the helix by breaking hydrogen bonds. Single stranded binding proteins keep the separated strands apart so that nucleotides can bind. Topoisomerase 2 helps relieve the strain on the helix. Initiation causes a replication fork to form.
Helicase
an enzyme formed from multiple polypeptides and doughnut in shape. The lagging strand passes through the centre of the enzyme while the leading strand passes on the outside, seperating the dna into two strands. ATP is needed to help the helicase move along the DNA molecule towards the replication fork (5' to 3') and break hydrogen bonds. It gets this from the cleaving of the 2 extra phosphate heads from a nucleotide.
Origin recognition complexes
bind to origins of initiation, allowing DNA Helicase to attach. Prokaryotes only have one replication fork, while eukaryotes have many to make replication faster and more efficient
DNA polymerase 3 (DNA replication)
an enzyme that cleaves off the extra 2 phosphate heads from nucleotides to create energy. DNA polymerase cannot initiate the process of DNA replication, there has to be a 3' end to which a free nucleotide can be added.
Elongation (DNA replication)
Synthesis of new DNA strands. RNA primase adds around 10-60 RNA nucleotides to the template strand (RNA primer), this will be removed later. DNA poly 3 adds free nucleotides (deoxynucleoside triphosphates) by cleaving off the heads. DNA poly 3 can only synthesize new DNA in a 5' to 3' direction towards the replication fork.
Laggiing strand (DNA replication)
synthesized discontinuously in short fragments away from the replication fork. Needs primers continuously at replication fork along the DNA parent strand and DNA poly 3 builds in short fragments called okazaki fragments. Eventually DNA poly 1 removes the RNA primer and fills it in with DNA bases. DNA ligase joins the fragments together. DNA poly 3 adds nucleotides moving away from the replication fork
Leading strand (DNA replication)
3' to 5' template is called the leading strand. Continuously built towards the replication fork
Termination (DNA replication)
as soon as the new strands are formed, they rewind automatically into their double helix shape. The two DNA molecules separate from each other and the replication machine (origin replication complex) is dismantled
What is Transcription?
The synthesis of RNA, using DNA as a template. it takes place in the nucleus, copying the gene sequence into the messenger RNA. It only occurs along one of the two strands of DNA, and genes can be transcribed repeatedly (happens all the time, vs DNA replication that only happens during cell division)
Promoter region
a DNA sequence near a gene that serves as the binding site of RNA polymerase (non-coding). The promoter however, is not transcribed. At the end of the promoter region there are several A-T bonds (TATA Box) because they are easy to break and it signals the RNA polymerase that it is reaching the end of the promoter. The binding of RNA poly to the promoter is mediated by transciption factors close to (proximal control) or far from (distal control) the promoter
Coding sequence
After the RNA polymerase has bound to the promoter, it causes the DNA strands to unwind and separate. This region that is transcribed by the RNA poly is called the coding sequence
terminator region
RNA polymerase will continue to tanscribe the DNA until it reaches a terminator sequence
Initiation (transcription)
The strand transcribed is called the antisense/template strand. The strand not transcribed is called the sense/coding strand, which has the same sequence as the mRNA with thymine instead of uracil. RNA Polymerase 2 binds to the DNA at a promoter sequence with the help of transcription factors (upstream of the gene). It unzips the DNA as it constructs an RNA transcript of the DNA. The binding site of RNA polymerase only recognizes the promoter region and can only bind upstream of it.
elongation (transcription)
RNA Polymerase builds the single stranded mRNA after it binds to the promoter in the 5' to 3' direction, by covalently bonding ribonucleotide triphosphates to create energy. When RNA polymerase leaves the promoter region another RNA polymerase can bind. Elongation starts as soon as RNA Poly binds to the promoter. If an error occurs, only one protein will be affected vs all of them in DNA. MRNA does not stay bonded to the DNA, it becomes displaced and the helix reforms
Termination (transcription)
RNA poly reaches the terminator sequence and dissociates with the template strand. The double helix of DNA reforms and the primary RNA transcript is produced (pre-mRNA) it is not ready to leave the nucleus just yet.
What is translation?
the process of protein synthesis in which the genetic information encoded in mRNA is translated into a sequence of amino acids in a polypeptide chain
sructure of ribosome
Translation takes place in free membrane bound ribosomes using the mRNA strand from transcription. Rbosomes are composed of two subunits (larhe and small) which are made of long strands of rRNA. The two subunits lock together when synthesizing a new protein with an mRNA trapped in the space between. The ribosome walks down the mRNA three nucleotides at a time to build the protein
A site
holds the tRNA carrying the next amino acid to be added to the polypeptide chain
P site
holds tRNA carrying the growing polypeptide chain
E site
site from which tRNA that has lost its amino acid is discharged
tRNA
Transfer RNA - contains an anticodon that base-pairs with a codon on the mRNA and has the corresponding amino acid attached to it according to the genetic code.
structure of tRNA
tRNA moves through three binding sites (A to P to E), then delivering amino acids to the growing polypeptide chain in translation. It picks up new amino acids when activated by a specific tRNA activating sequence, there are 20 of these that correspond to the 20 amino acids. This costs ATP. tRNA has a complementary anticodon for the 20 seuqences, and the energy in the bond linking the tRNA to the amino acid is used in translation to form peptide bonds between amino acids.
codons
there are 64 different codon combinations, codons are a triplet of bases that codes for amino acids. The coding rehion always starts with a START codon (AUG), and the coding region of mRNA terminates with a stop codon (UAA, UAG or UGA), causing the release of the polypeptide
initiation (translation)
the 5' end of the mRNA binds to the small subunit of the ribosome. The small subunit of the ribosome moves along the mRNA molecule in a 5' to 3' direction until it reaches the start codon. A molecule of tRNA complementary to the start codon (UAC) binds to the p site, and the large subunit of the ribosome binds to it as well as the small subunit
elongation (translation)
a second tRNA (with amino acid) complementary to the second codon on the mRNA binds to the A site of the ribosome. The amino acid carried by the first tRNA is transferred to the second, causing the polypeptide to increase in length and the dipeptide being attached to the second tRNA. The ribosome then moves one codon along the mRNA and the first tRNA is moved to the E site, the second moving to the P site. Another tRNA binds that is complementary to the next codon and the process keeps repeating.
termination (translation)
when a stop codon is reached at the A site translation is stopped. A protein known as the release factor then fills the A site. It helps to break the bond linking the tRNA in the P site with the polypeptide chain. This causes the ribosome subunits to dissassemble, releasing the mRNA and the protein. The new protein is then folded and modified to the targeted areas of the cell where it is required
modification of polypeptides into their functional state
after translation, polypeptides are modified so they are fully functional. This can happen through changes to the side chain of amino acids, folding of the polypeptides, removing part of the polypeptide chain, combining polypeptides (quarternary structure), or forming conjugated proteins by adding a non-polypeptide component
methylation
the addition of methyl groups to the DNA (specifically cytosine), it helps to inhibit transcription, and bind the DNA more tightly to the histone to make it less accessible to transcription factors (in the promoter region)
acetylation
the addition of acetyl groups to histones to promote transcription. It does this by binding the DNA more loosely to make it more accessible to transcription factors
methylation of histones
Helps promote and inhibit transcription. Whether or not a gene is expressed depends on where it is methylated. Methylation of lysine 4 (near the tip) causes transcription, and methylation of lysine 9 or 27 (along the tail) causes the gene to be silenced
Epigenetics inheritance through changes to gene expression
Epigenetics allows a genome to be applied different ways in different situations. Methylation patterns can be influenced by experiences (can increase likelihood of certain diseases and behaviours), these can be reversible, but some tags are passed down to the next generation
factors contributing to methylation patterns
Human hair and skin colour can be affected by temperature and sunlight exposure, pollution can affect the expression of proteins that regulate the immune system, air pollution is most damaging during pregnancy. As well as the environment of a cell
methylation of monozygotic twins
DNA methylation patterns will differ between twins and continue to diverge over time as a consequence of environmental exposure. DNA methylation patterns in twins can be used to identify genes involved in the development of specific diseases that are present in a single twin. Variation increases in the levels of hypermethylation (high levels) and hypomethylation (low levels) with age.
Hershey chase (viruses pt 1)
Viruses are made up of proteins and nucleic acid (allowed them to figure out which was the genetic material), they inject their genetic material into a host cell to reproduce. Amino acids containing radioactive isotopes to label the virus. Sulfur for the protein coat, and phosphorus for the DNA. The bacteriophage was then combined with E-coli bacteria. When separate with a centrefuge the virus remained a liquid and the bacteria formed a pellet.
Hershey chase (tracking protein pt 2)
The viral proteins in the bacteriophage were tagged with sulfur. A blender was then used to separate the infected bacteria and extra virus parts. Then a centrefuge was used, being more dense, the bacteria cells were isolated because they formed a pellet at the bottom of the tube. Radioactive protein molecules were not present in the bacteria cells.