Biomed sciences I exam# 1

Describe the structure of DNA

• Double Helix

• Antiparallel

• Complementary

Monomeric units of DNA

nucleotides

Which Nitrogenous bases contains Adenine (A) and Guanine (B)

Purines

Which Nitrogenous bases contain Cytosine (C) and Thymine (T)

Pyrimidines

What 3 things make a nucleotide

• Nitrogenous base

• Sugar

• Phosphate

What are Sugars

deoxyribose (pentose)

What is the phosphate backbone?

phosphoric acid

What facilitates the formation of polynucleotides

Phosphodiester bonds between 3’ - OH of one sugar and 5’ - OH of next sugar

Each DNA molecule is formed by 2 ______ joined by ______ between bases

polynucleotide chains, hydrogen bonds

What can heat/alkali destruction of hydrogen bonds cause?

DNA denaturation (the 2 strands separate, since phosphodiester bonds are resistant)

Dissociation is reversible by reversal of denaturing conditions, this is called ______?

renaturation (reannealing)

What is the Melting temperature (Tm)?

temperature at which 50% of DNA is single stranded

What is Hybridization?

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

What are the two types of DNA?

Mitochondrial and Nucleic

What are the two properties of Genetic information

- Inherited (replication)

- Expressed (transcription & translation)

What shape is Mitochondrial DNA?

Circular, double-stranded (endosymbiotic theory)

how much of total cellular DNA does Mitochondrial DNA make up?

1%

Mitochondrial DNA lacks _____ and is _____ rich

histones, GC

How many genes does Mitochondrial DNA have?

37 genes

• 13 for energy production

• 24 for rRNA and tRNA

Mitochondrial DNA is derived exclusively from the _______

Mother

Why is Mitochondrial DNA a good marker for human ancestry?

there is no recombination

what percent of Nucleic DNA is non-coding?

98%

How long is the Nucleic DNA

6ft/cell, encoding for 22000 genes

What is the Centromere?

constricted region of the chromosome

What is the Attachment site for the spindle microtubules and required for correct mitosis

Centromere

What are Telomeres?

Caps at the ends of chromosomes

What chromosome structure has Repeats of TTAAGGG (1000x) and is shortened with each cell division?

Telomeres

What is the short arm of the chromosome?

p

what is the long arm of the chromosome?

q

Super-compacted DNA-protein complexes that chromosomes are formed by

Chromatin

Characteristics of Euchromatin

• Less condensed

• Located on chromosome arms

• Unique sequences

• Many genes

• Transcribes often

Characteristics of Heterochromatin

• More condensed

• Located at centromeres, telomeres, and other specific places

• Repeated sequences

• Few genes

• infrequent transcription

Histone octamer core (2 molecules of each H2A, H2B, H3, H4) + dsDNA wrapped 2x around the core makes what?

Nucleosome

What connects adjacent nucleosomes (beads on a string)?

Linker DNA + histone H1

What do supercoiled nucleosomes make?

chromatin fibers

What does the compaction of chromatin fibers make?

chromosomes

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

Cell cycle

Which phase does DNA replication take place?

S phase

Which phase does DNA repair take place?

G2 phase

How long does genome duplication take in a cell?

9 hours

DNA replication begins at specific sequences called?

Origins of replication

• 100/chromosome in eukaryotes, only 1 in prokaryotes

DNA replication is ______ (each new dsDNA contains one parental and one daughter strand) and _____ (in both directions from the origin of replication)

Semiconservative and bidirectional

What is the Y-shaped region where synthesis occurs?

Replication fork

What is Helicase necessary for?

Breaks hydrogen bonds and separates the 2 strands

What is DNA Topoisomerase necessary for?

Relaxes coils and unwinds DNA ahead of replication fork

What are Single-stranded binding proteins (SSBP) necessary for?

Attach to each strand to prevent reannealing

What is Primase (Subunit of DNA polymerase) necessary for?

Synthesizes an RNA primer (approx. 15nt) by copying the parental strand

What is DNA polymerase necessary for?

• Adds dNTPs to the free 3’-OH of the RNA primers (Always leaves a free 3’-OH to accept more dNTPs)

DNA polymerase Reads parental strand in the ______ direction, synthesizes daughter DNA in the ______ direction, and has ______ direction exonuclease activity to remove the RNA primer

3’→5’, 5’→3, 5’→3’

Which proofreading activity excises mismatched nucleotides, which allows for normal continuation of synthesis

3’→5’ exonuclease activity

Polymerase (alpha) function and proofreading capability?

Function: Contains primase, initiates DNA synthesis

Proofreading: (-)

Polymerase (beta) function and proofreading capability?

Function: Repair

Proofreading: (-)

Polymerase (delta) function and proofreading capability

Function: Elongates Okazaki fragments of the lagging strand

Proofreading: (+)

Polymerase (epsilon) function and proofreading capability

Function: Elongates the leading strand

Proofreading: (+)

Polymerase (gamma) function and proofreading capability

Function: Replicates mitochondrial DNA

Proofreading: (+)

What is DNA ligase necessary for?

Joins the free 3’-OH group and the free 5’-phosphate group to form a phosphodiester bond

The leading strand runs ______ toward the replication fork and is replicated continuously from an RNA primer by ________

3’→5’, DNA polymerase

What direction does the Lagging strand run away from the replication fork?

3’→5’

Lagging strand is replicated discontinuously from multiple RNA primers by ______ to form ________

DNA polymerase, Okazaki fragments

Lagging strand primers are removed by ______ and gaps are filled

5’→3’ exonuclease

What are ends of adjacent fragments in lagging strand joined by?

DNA ligase

What does the removal of RNA primer form the 3’-end of the daughter strand during lagging strand synthesis lead to?

Shorter newly synthesized DNA strand

What is the main function of Telomerase?

Preserve the length of the telomeres by copying a small number of repeats, after which the complex moves forward to the 3’-end of the overhang and repeats the process

Do somatic cells express telomerase?

No

In what cells is telomerase expressed in?

germline cells and cancer cells

Is Telomerase a ribonucleoprotein?

Yes

Why is telomerase considered a ribonucleoprotein?

because it has the enzyme Reverse transcriptase (TERT gene) and the RNA template (TERC gene)

Examples of Spontaneous DNA damage

- Alkylation

- Depurination

- Deamination

Types of DNA damage

• Replication errors

• Spontaneous

• Mutagen-induced damage

Fidelity of replication errors occur approx. 1 in every _____ nucleotide

10^9 - 10^10

Mismatched bases during replication is an example of _____ damage

endogenous

DNA damage by mutagens is an example of _____ damage

exogenous

What are permanent changes (undetected/unrepaired)

mutations

are errors and mutations the same?

No

Endogenous Mutagens ex.

Free radicals

Exogenous Mutagens ex.

• Alkylating agents

• Radiation (ionizing, X-rays, UV)

• Chemicals (pollutants, pesticides, plant and microbial toxins, smoking, chemotherapeutics)

Mutagens that induce normal cells to become cancerous are called?

Carcinogens

Types of Single strand damage repair

Excision repair

• Base excision repair (BER)

• Mismatch repair (MMR)

• Nucleotide exision repair (NER)

Types of Double-strand damage repair

Recombination repair

• Homologous recombination (HR)

• Non-homologous end joining (NHEJ)

Generic DNA excision repair pathway

• Detection of damage

• Excision of damaged DNA by endo-/exonucleases

• Replacement of excised region by DNA polymerase

• Nick sealing by DNA ligase

What is the simplest direct damage reversal mechanism?

repair done without cuts in the DNA backbone

Base excision repair mechanism

Repair small distortions in DNA caused by damaged bases

Can base excision repair happen spontaneously?

Yes

• Depurination (>1000/day)

• Deamination (C→U) (>100/day)

  • Deamination of methylated C→T escapes repair

Does the sugar-phosphate backbone remain intact during Base excision repair?

YES!

How does Mismatch repair (MMR) work

Removes and repairs mispaired, but undamaged bases added during replication (escape DNA polymerase proofreading)

Mismatch repair (MMR) corrects small _________

insertions or deletions

Process of MMR

• Mismatched bases in newly synthesized DNA produce distortions and trigger MMR

• Newly synthesized strand recognized by presence of nicks

• Parental strand used as template

Defects in MMR can lead to what syndrome?

Lynch syndrome

What does Nucleotide excision repair (NER) do?

Detects and corrects bulky damage that distort the DNA double helix → replication block

the two major types of damage fixed by Nucleotide excision repair (NER)

• Cigarette smoke - benzo(a)pyrene-guanine adducts

• UV radiation - adjacent pyrimidine dimers (T-T)

  • Unresolved T-T dimers result in UV-induced skin cancers

Defects In NER results in what disorder?

xeroderma pigmentosum

What induces double-strand breaks in the DNA?

Ionizing radiation, ROS, and inadvertent action of nuclear enzymes

What is the main cause of genomic instability and large mutations?

Double-strand breaks in DNA

Describe homologous recombination process

Information from the sister chromatid is used as template to replace the missing region

What phase of the cell cycle does homologous recombination mostly occur in

S-phase

Why is Homologous recombination the preferred mechanism in yeast?

Lots of repetitive sequences in animal DNA → gene amplification

One advantage and disadvantage of homologous recombination

Advantage: More precise

Disadvantage: More complex

What is the primary method of DSB repair in mammalian cells?

Non-homologous end-joining

When does non-homologous end-joining mostly occur?

G1 phase of cell cycle

What does non-homologous end joining generally result in?

Deletions or small insertions