DEOXYRIBUNUCLEIC ACID (DNA)

• DNA mutation

• DNA maturation

Other terms for DNA Replication

DNA Replication

A process in which a DNA creates an identical copy of itself.

Semiconservative replication mechanism.

DNA replication follows this mechanism.

• 2 rounds

• 4 daughter cells

• How many rounds occur in DNA replication?

• How many daughter cells does it produce?

• 2 daughter strands with one old and one new strand

• 2 daughter strands with two new strands

The four daughter strands in DNA replication consists of?

Meselson and Stahl

Who proposed the semiconservative nature of DNA replication?

Semiconservative Replication Mechanism

It suggests that every time a new DNA is synthesized, there is always one parent strand and one daughter strand.

• Initiation

• Elongation

• Termination

What are the three phases of DNA replication?

Origins of replication

DNA Replication starts at specialized sites called?

Replication bubble

Once the origins of replication begins, it will form a?

Replication forks

Each replication bubble contains?

Replication forks

This is where the DNA unwinds.

Replication bubble

This is the space between the replication forks.

Replisomes

Once DNA is opened, replication machinery (enzymes and proteins) are loaded to the single strand, which will form the templates for the daughter strand.

Replisomes

These are a set of proteins and enzymes that participates in the copying of the DNA.

Elongation

Once the replication machinery is in place, and the DNA has been opened up, replication enters what phase?

The very first sequence

Elongation phase begins when this is synthesized.

• Leading strand

• Lagging strand

• The __ is copied in a continuous manner

• The __ is copied in a discontinuous manner

3’ to 5’

The leading strand and lagging strand are read in what direction?

5’ to 3’

Leading strand and lagging strand synthesis occurs in what direction?

Termination

This occurs when the two replication forks moving in opposite directions meet, and the replication complexes are disassembled.

Termination

What phase?

The replisome is removed from the DNA and the strands are glued together by a specific enzyme.

Helicase

Harnesses chemical energy from ATP hydrolysis to separate DNA strands at the replication fork.

Single-Strand DNA Binding Protein

They bind selectively to single-stranded DNA (ssDNA) as soon as it forms, stabilizing and protecting it.

Supercoiling

As the helicase unwinds DNA, the twist ahead of the fork increases in tension, and results to?

Positive supercoiling

It condenses DNA

Negative supercoiling

It relaxes DNA and begins to separate.

DNA Topoisomerase I

It relaxes supercoiled structures.

DNA Topoisomerase II

It introduces negative supercoils through coupling to ATP hydrolysis.

DNA gyrase

Bacterial topoisomerase

Primase

It has the role of synthesizing RNA primers

RNA Primers

These are the starting material for DNA replication.

The first to be placed in the new strand.

RNA Primers

It acts as a foundation of DNA replication.

Free 3’ end

RNA primers are section of nucleid acid with a?

False. Primase is removed at the end of replication.

True or False. Primase is retained until transcription phase.

DNA Polymerase

The most important enzyme in DNA Replication

DNA Polymerase

It is responsible for copying DNA during replication.

DNA Polymerase III

A prokaryotic enzyme that directs the synthesis of a new DNA strand starting from the RNA primer.

DNA Polymerase I

A prokaryotic enzyme that functions as an exonuclease by removing RNA primer and replaces it with deoxynucleotides.

• RNA primer

• Deoxynucleotides

DNA Polymerase I It functions as an exonuclease by removing __ and replaces it with __.

Exonuclease

These are enzymes that can cut a certain portion of a nucleic acid.

• High fidelity replication

• The exonuclease double checks for errors by Pol III / checks the correctness

• Enzymes with exonucleases participates in maintaining what kind of replication?

• What does it mean?

DNA Polymerase alpha

A eukaryotic enzyme that acts as an initiator polymerase. It begins the DNA replication process, but does not finish the job.

DNA Polymerase delta and epsilon

Delta - lagging strand

Epsilon - leading strand

It directs the synthesis of a new DNA strand from lagging strand (_) and leading strand (_).

DNA Polymerase Delta and Epsilon

Eukaryotic polymerase that have exonuclease activity.

DNA Ligase

It seals nicks and breaks in dsDNA molecules through the formation of phosphodiester bonds between existing DNA strands.

DNA Ligase

It is utilized in joining Okazaki fragments.

Energy dependent

Is DNA Ligase energy-dependent or non-energy-dependent?

Bidirectional Replication

A replication pattern that is common among eukaryotes.

Rolling Circle Replication

A replication pattern that is common among bacteria and bacteriophage.

At least 50 nucleotides are lost in telomeres

DNA polymerase is not capable to completely copy DNA at the very ends of chromosomes, which results to __ nucleotides lost during each cell cycle.

The loss of 50 nucleotides in telomeres during each cycle.

The reason for cell aging.

Telomerase

An enzyme that maintains the length of telomeres by addition of guanine-rich repetitive sequence.

Guanine-rich repetitive sequence

Telomerase maintains the length of telomeres by adding?

• Gametes

• Stem cells

• Tumor cells

Increased telomerase activity is seen in?

Cancer cells

A case where the telomere’s length is the same, which is also referred as immortal cells.

HeLa cell

An example of immortal cancer cell that came from human cervical cancer cells.

It is a viral culture media.

Mitochondrial DNA

It is a type of DNA that is inherited from mothers only.

Mitochondrial DNA

» circular pattern, and same components

It has a similar organization with bacterial genome.

Stochastic

Mitochondrial DNA replication is __, meaning there is no control over which particular copies are replicated.

Strand displacement model

Mitochondrial DNA replication follows the ___, wherein the replication is unidirectional around the circle and there is no replication fork for each strand.

DNA Polymerase Gamma

Mitochondrial DNA replication utilizes this as its primary enzyme.

Mutations

It results from changes in the nucleotide sequence of DNA or from deletions, insertions, or rearrangements of DNA sequences in the genome.

Evolution from Neanderthals to Homo sapiens

An example of a best beneficial mutation.

• Genetic disorders

• Cancers

Examples of Bad Mutations

• Induced mutation

• Spontaneous mutation

KINDS OF MUTATIONS.

• Occurs as a result of interaction of DNA with an outside agent or mutagen, which causes DNA damage (external factors).

• Occurs as a result of natural processes in cells.

• UV rays

• X-rays

• Radioisotopes

• Benzene

Example of external factors that cause induced mutations

Aplastic anemia

What disease can you get from benzene?

• Point mutation

• Insertions and Deletions (InDels)

Types of DNA Mutation

Transition and Transversion Mutations

• Silent mutations

• Missense mutations

• Nonsense mutations

• Describe point mutations.

• What are the 3 types?

Silent mutation

What kind of mutation?

There is a change in the sequence but the encoded amino acid is the same.

Missence mutations

What kind of mutation?

There is a change in the nucleotide sequence and there is a change in the amino acid encoded.

Nonsense mutations

What kind of mutation?

There is a change in the sequence and resulted to a stop codon.

A serious type of mutation.

Frameshift mutation

What kind of mutation?

A nucleotide is inserted in an original sequence, which is now included in the reading.

Point mutations

Also known as nucleotide substitution, in which a base pairs with an inappropriate partner during DNA replication.

Transition mutation

One purine (or pyrimidine) is replaced with another.

Transversion Mutations

A purine is substituted for a pyrimidine, or vice versa.

Insertions and Deletions

» Frameshift mutation

It refers to the addition and removal of one or more base pairs, respectively. It results to “shifts” in the reading frame of the codons.

DNA damage

It is a chemical alteration to DNA which can be introduced by many different ways and if left unrepaired, may lead to an induced DNA mutation.

• Single base changes

• Structural distortion

• DNA backbone damage

3 Major Classes of DNA Damage

DNA backbone damage

It includes the formation of abasic sites, single and double-strand DNA breaks. It is the most dangerous DNA damage.

1. Single base changes (conversion)

2. Structural distortion

DNA DAMAGE.

1. Affects the DNA sequence but has only a minor effect on overall structure.

2. Primarily affects the overall structure that may impede transcription and replication by blocking the movement of polymerases.

1. Oxidation

2. Deamination

3. Alkylation

DNA Damage: Single Base Changes

1. Caused by potent oxidizing agents. Reactive oxygen species can generate 8-oxoguanine (oxoG), a highly mutagenic damaged guanine.

2. Replacement of the amino group of cytosine with oxygen converts cytosine to uracil, a base that should be present only in an RNA chain.

3. Caused by alkylating agents. Electrophiles attack negatively-charged DNA molecules and add alkyl groups.

1. Intercalating Agents

2. Base analogs

3. UV radiation

DNA Damage: Structural Distortion

1. Contain polycyclic rings, which inserts between DNA bases resulting to the distortion of the DNA helix.

2. Mutagenic chemicals that substitute for normal bases.

3. Cross-links pyrimidines on the same DNA strand, commonly forming pyrimidine dimers. Majority of pyrimidine dimers form a cyclobutyl ring, while others do not.

DNA Repair Systems

These are responses of biological system to different types of DNA damage.

• Damage Reversal

• Damage Removal

2 categories of DNA Repair

• DNA Photolyase

• DNa Methyltransferase

Two DNA repair enzymes capable of reversing DNA damage

DNA Photolyase

It reverses the pyrimidine dimers caused by UV radiation through the process called photoreactivation or “light repair”.

DNA Methyltransferase

It reverses the DNA damage caused by O6-methylguanine by catalyzing the removal of the methyl group from the damaged guanine.

Damage Removal

It refers to the repair of both single base changes and structural distortion.

• Base Excision Repair, Mismatch Repair

• Nucleotide Excision Repair

• Two main pathways for repair of single base changes

• Repair of structural distortion

Base Excision Repair

It is initiated by a group of enzymes called DNA glycosylases, which breaks the glycosidic bond between the damaged base and its sugar, leaving an AP site.

Mismatch repair

It corrects mistakes that occur during DNA replication and that are not proofread by the DNA polymerase.

Double-strand breaks

Out of the various types of DNA damage, ___ are the most harmful to cells and are often linked to cell death or cancer.

• Homologous recombination

• Nonhomologous end-joining

Double strand breaks are repaired by?

Homologous Recombination

It repairs double-strand breaks by retrieving genetic information from an undamaged homologous chromosome.

Nonhomologous end-joining

It occurs simply by the rejoining of broken DNA ends and does not require another template duplex such as sister chromatid (does not utilize homologous copy).