Biomed 1 Lecture 1- DNA Organization, Replication, and Repair

What is the overall shape of the DNA molecule?

Double helix

Are the two strands of DNA parallel or antiparallel?

Antiparallel (run in opposite directions, 5' to 3')

What does it mean that DNA strands are complementary?

Each base on one strand pairs specifically with a base on the other strand (A-T, G-C)

What are the monomeric units of DNA called?

Nucleotides

What three components make up a nucleotide?

A nitrogenous base + a sugar (deoxyribose) + a phosphate group

What are the two purine bases in DNA?

Adenine (A) and Guanine (G)

What are the two pyrimidine bases in DNA?

Cytosine (C) and Thymine (T)

What sugar is found in DNA nucleotides?

Deoxyribose (a pentose sugar)

What forms the phosphate backbone of DNA?

Phosphoric acid, linked via phosphodiester bonds

What type of bond links the 3'-OH of one sugar to the 5'-OH of the next sugar in DNA?

Phosphodiester bond

What do phosphodiester bonds link together to form?

Polynucleotides

How many polynucleotide chains form one DNA molecule?

Two, joined by hydrogen bonds between bases

How many hydrogen bonds form between A and T?

2 hydrogen bonds

How many hydrogen bonds form between G and C?

3 hydrogen bonds

What is DNA denaturation?

Heat or alkali destruction of hydrogen bonds, causing the two DNA strands to separate (phosphodiester bonds remain intact/resistant)

What is DNA renaturation (reannealing)?

The reversible reassociation of separated DNA strands after reversal of denaturing conditions

What is the melting temperature (Tm) of DNA?

The temperature at which 50% of the DNA is single-stranded

What determines the melting temperature (Tm) of a DNA molecule?

Its base composition (higher GC content = higher Tm)

What is hybridization (in the context of DNA)?

The tendency of single-stranded DNA to bind complementary sequences of DNA or RNA

What are the two locations of DNA within a eukaryotic cell?

Mitochondrial DNA and nuclear (nucleic) DNA

What is the shape of mitochondrial DNA?

Circular and double-stranded

What theory explains the circular, double-stranded nature of mitochondrial DNA?

Endosymbiotic theory

What percentage of total cellular DNA is mitochondrial DNA?

Approximately 1%

Does mitochondrial DNA have histones?

No, it lacks histones

Is mitochondrial DNA GC-rich or AT-rich?

GC rich

How many genes does mitochondrial DNA contain?

37 genes

How many mitochondrial genes encode proteins for energy production?

13 genes

How many mitochondrial genes encode rRNA and tRNA?

24 genes

From which parent is mitochondrial DNA inherited?

Exclusively from the mother

Why is mitochondrial DNA useful as a marker for human ancestry?

Because it undergoes no recombination

What percentage of nuclear (nucleic) DNA is non-coding?

Approximately 98%

Approximately how long is the nuclear DNA per cell if stretched out?

Approximately 6 feet per cell

Approximately how many genes does nuclear DNA encode?

Approximately 22,000 genes

What is a centromere?

The constricted region of a chromosome that serves as the attachment site for spindle microtubules and is required for correct mitosis

What are telomeres?

Caps at the ends of chromosomes

What repeat sequence makes up telomeres, and how many times is it repeated?

Repeats of TTAGGG, about 1000 times

What happens to telomeres with each cell division?

They are shortened

What is chromatin?

Super-compacted DNA-protein complexes that form chromosomes

Is euchromatin more or less condensed than heterochromatin?

Less condensed

Where is euchromatin located on the chromosome?

On chromosome arms

What type of DNA sequences does euchromatin contain?

Unique sequences

Does euchromatin contain many or few genes?

Many genes

How often is euchromatin transcribed?

Often (actively transcribed)

Where is heterochromatin located on the chromosome?

At centromeres, telomeres, and other specific places

What type of DNA sequences does heterochromatin contain?

Repeated sequences

Does heterochromatin contain many or few genes?

Few genes

How often is heterochromatin transcribed?

Infrequently

What is a nucleosome?

The structural unit of chromatin, consisting of a histone octamer core plus DNA wrapped around it

What proteins make up the histone octamer core of a nucleosome?

Two molecules each of H2A, H2B, H3, and H4

How many times does DNA wrap around the histone octamer core?

Twice (1.65-2 times)

What connects adjacent nucleosomes to form the "beads-on-a-string" structure?

Linker DNA plus histone H1

What is formed when nucleosomes are supercoiled?

Chromatin fibers (solenoid, tubular coils)

What is formed by the compaction of chromatin fibers?

Chromosomes

What two cellular processes require DNA synthesis?

Replication and repair

What is the cell cycle?

The sequence of ordered events during which proliferating cells replicate their chromosomes and separate into daughter cells

During which phase of the cell cycle does DNA replication occur?

S phase

During which phase of the cell cycle does DNA repair occur?

G2 phase

Approximately how long does genome duplication take in a cell?

Approximately 9 hours

What are the specific DNA sequences where replication begins called?

Origins of replication (ori)

Approximately how many origins of replication exist per chromosome in eukaryotes?

Approximately 100 per chromosome

Approximately how many origins of replication exist in prokaryotes?

Only 1

What does it mean that DNA replication is "semiconservative"?

Each new double-stranded DNA molecule contains one parental strand and one newly synthesized (daughter) strand

What does it mean that DNA replication is "bidirectional"?

Replication proceeds in both directions from the origin of replication

What is a replication fork?

The Y-shaped region of DNA where synthesis occurs

What is the function of helicase in DNA replication?

It breaks hydrogen bonds and separates the two DNA strands

What is the function of DNA topoisomerase in DNA replication?

It relaxes coils and unwinds DNA ahead of the replication fork

What is the function of single-stranded binding proteins (SSBPs)?

They attach to each separated DNA strand to prevent reannealing

What is primase and what does it do?

A subunit of DNA polymerase that synthesizes an RNA primer (approximately 15 nucleotides) by copying the parental strand

In what direction does DNA polymerase read the parental (template) strand?

3' to 5'

In what direction does DNA polymerase synthesize the new (daughter) DNA strand?

5' to 3'

To what part of the RNA primer does DNA polymerase add dNTPs?

The free 3'-OH end

What exonuclease activity does DNA polymerase use to remove the RNA primer?

5' to 3' exonuclease activity

What exonuclease activity does DNA polymerase use for proofreading (removing mismatched nucleotides)?

3' to 5' exonuclease activity

What does the proofreading (3' to 5' exonuclease) activity of DNA polymerase do?

It excises mismatched nucleotides, allowing normal continuation of synthesis

Which DNA polymerase contains primase and initiates DNA synthesis?

DNA polymerase alpha (does NOT have proofreading activity)

Which DNA polymerase is responsible for DNA repair?

DNA polymerase beta (does NOT have proofreading activity)

Which DNA polymerase elongates the Okazaki fragments of the lagging strand?

DNA polymerase delta (HAS proofreading activity)

Which DNA polymerase elongates the leading strand?

DNA polymerase epsilon (HAS proofreading activity)

Which DNA polymerase replicates mitochondrial DNA?

DNA polymerase gamma (HAS proofreading activity)

What is the function of DNA ligase?

It joins the free 3'-OH group and the free 5'-phosphate group to form a phosphodiester bond, sealing nicks in the DNA backbone

In what direction does the leading strand run relative to the replication fork?

It runs 3' to 5' toward the replication fork

How is the leading strand replicated?

Continuously, from a single RNA primer, by DNA polymerase

In what direction does the lagging strand run relative to the replication fork?

It runs 3' to 5' away from the replication fork

How is the lagging strand replicated?

Discontinuously, from multiple RNA primers, by DNA polymerase, forming Okazaki fragments

Approximately how many nucleotides long is an Okazaki fragment?

Approximately 200 nucleotides

How are RNA primers removed from Okazaki fragments and the resulting gaps filled?

Primers are removed by 5' to 3' exonuclease activity and gaps are filled by DNA polymerase

How are the ends of adjacent Okazaki fragments joined?

By DNA ligase

What is the "end-replication problem"?

Removal of the RNA primer from the 3' end of the daughter strand during lagging strand synthesis leads to a shorter newly synthesized DNA strand

What is the long-term consequence of the end-replication problem over multiple rounds of replication?

Progressive shortening of the telomeres

What type of molecule is telomerase?

A ribonucleoprotein

What gene encodes the reverse transcriptase component of telomerase?

TERT gene

What gene encodes the RNA template component of telomerase?

TERC gene

Approximately how large is the hexanucleotide repeat region used as the telomerase RNA template?

3-20 kb (repeat sequence 3'-AAUCCC-5')

How does telomerase preserve telomere length?

By copying a small number of repeats using its RNA template, then translocating to the 3' end of the overhang and repeating the process

Do somatic cells normally express telomerase?

No, somatic cells do not express telomerase, leading to progressive chromosome shortening and cell aging

Which cell types express telomerase?

Germline cells and cancer cells

What causes replication errors during DNA synthesis?

Failure of the proofreading activity of DNA polymerase to detect and repair mismatches

What is the approximate fidelity (error rate) of DNA replication?

Approximately 1 error in every 10^9 to 10^10 nucleotides

What are the three main types of spontaneous DNA damage?

Alkylation, depurination, and deamination

What are the two broad categories of DNA damage sources?

Spontaneous damage and mutagen-induced damage