DNA replication, expression, traslation Flashcards
Cell Cycle
To divide, a cell must complete several important task:
- grow
- copy its genetic material (DNA)
- physically split into 2 daughter cells
The cell cycle is divided into 2 major phases:
1) INTERPHASE
2) MITOTIC (M) PHASE
Interphase
phases:
- G1
cellular contents, excluding the chromosomes, are duplicated.
- S
each of the 46 chromosomes is duplicated by. the cell.
- G2
the cell “double checks” the duplicated chromosomes for error, making any needed repairs
the cell grows and makes a copy of its DNA.
Mitotic (M) Phase
the cell separates its DNA into two sets and divides its cytoplasm, forming 2 new cells.
At each cell division, a cell must copy its genome. It is essential that this process occurs accurately.
DNA acts as a template for its own replication -> SEMI-CONSERVATIVE REPLICATION.
Enzymes
- DNA Helicase
unwinds the DNA - DNA Polymerase
synthesis DNA 5’-3’ direction - DNA Topoisomerase
relieves the tension in DNA - DNA Primase
synthesis RNA primers - Ribonucleases
degrades RNA primers - DNA Ligase
joins DNA fragments - Telomerase
replicates the ends of the chromosome
DNA unwinding
DNA helicase:
- unwinds the DNA
- uses ATP to propel itself along the DNA
Single-strand DNA binding protein:
binds and keep the strands apart
DNA topoisomerase:
relives the tension
DNA synthesis
- DNA polymerase requires a template
- DNA polymerase requires a primer
short segment of RNA complementary to the template with a 3’ OH.
a new strand of DNA is always synthesised in a 5’ to 3’ direction. -> it elongates from a free 3’OH.
Replication Fork
DNA replication progresses 5’-3’ so replication on the leading strand is continuous.
As DNA replication cannot progress in the opposite direction, replication on the lagging strand is discontinuous.
The short DNA sequences synthesised on the lagging strand are known as OKAZAKI FRAGMENTS.
Sliding Clamp
the DNA polymerase remains attached to the DNA template by interaction with a protein called SLIDING CLAMP.
A new clamp has to be loaded on the lagging strand as each Okazaki fragment is synthesised.
Lagging Strad
- DNA primate attached RNA to template
- DNA polymerase III adds nucleotides until it reaches the previous primer
- RNAse H digests the RNA primer, leaving a gap.
- DNA polymerase I fills in the gaps.
- DNA ligase then joins the fragments together
Telomeres
repetitive regions at the very ends of chromosomes are called TELOMERES.
Telomeres act as CAPS that protect the internal regions of the chromosomes, and they are worn down a small amount in each round of DNA replication.
They are G-rich series of repeat bases (TTAGGG repeated hundreds or even thousands of times in mammals).
Telomeres: Problem
In lagging strand Okazaki fragments cannot cover the end of the chromosomes.
There’s no way to get the fragments started because the primer would fall beyond the chromosome end.
Part of the DNA at the end of a eukaryotic chromosome goes uncopied in each round of replication.
This leaves a SINGLE-STRANDED OVERHANG. Over multiple rounds of cell division, the chromosomes will get shorter and shorter as this process repeats.
Telomeres: Solution
some cells have an enzyme called TELOMERASE which is an RNA-dependent DNA polymerase, meaning an enzyme that can make a DNA using RNA as a template.
The enzymes binds to a special RNA molecule that contains the sequence complementary to the telomeric repeat. Telomerase recognises the tip of an existing repeat sequence and uses the RNA template within the enzyme to add additional repeats to the telomere DNA.
When the overhang is long enough, a matching strand can be made by DNA polymerase alfa, which has its own primase submit -> so doesn’t need a primer.
Fidelity and Proofreading
DNA polymerase makes a mistake once every 107 nucleotide pairs it copies.
Detects incorrect base-pairing -> active site geometry accommodates only A-T & G-C base pairs -> proofreading: 3’ to 5’ exonuclease activity.
Despite this process, DNA can still be damaged, as a result of:
- UV Light
2 adjacent thymine bases become covalently attached to each other. Leads to stalling of the replication machiner. Failure to repair thymidine dimer is the problem in XERODERMA PIGMENTOSUM.
- Ionising radiation
- toxic chemical agents
- reactive oxygen species
DNA damage can result in mutations in genes that can lead to altered coding for proteins resulting in LOSS or GAIN OF FUNCTIONS.
Mutation: GAIN OF FUNCTION
a DNA sequence change that leads to increased or alternative activity.
Like:
- overactivity of a gene product overrides existing control mechanism leading to a cancerous cell.
- the amino acid change leads to a change in a protein’s interaction with a inhibitory ligand.
Mutation: LOSS OF FUNCTION
a DNA sequence change that leads to a decreased activity.
Like:
- the nucleotide change leads to a loss of expression of the protein
- the amino acid change in a protein’s interaction with its ligands.
Point Mutations
Genome DNA is transcribed to messanger RNA which is then translated into an amino acid sequence.
Every 3 nucleotides on the mRNA, starting with a specific start sequence (AUG) , is called a CODON which corresponds with a specific amino acid.
Substituting a nucleotide can alter the codon in a number of ways:
- Silent
- Missense
- Nonsense
Silent Mutation
is when a nucleotide substitution results in a different codon that still encodes the same amino acid.
Therefore, the protein in unaffected in function and the phenotype of the organism is not simnifically altered.
Missense Mutation
is when a nucleotide substitution results in a codon that encodes a different amino acid.
Therefore, the primary protein sequence is altered which may be conservative or radical:
- CONSERVATIVE substitution
similar amino acid R group size and charge, similar protein shape and function.
- RADICAL substitution
new amino acid R group different in charge or size, protein may have altered secondary or tertiary structure affecting function.
Nonsense Mutation
is when a nucleotide substitution results in a stop codon that stops translation.
Therefore, the protein is truncated and may not function properly or even at all.
Protein Truncation:
a process during translation that detects transcripts with premature stop codons and decades them.
