B: Master document

Question 1

What is the difference between a purine and pyrimidine?

Answer 1

A pyrimidine is a heterocylic aromatic ring like benzene which a nitrogen atom in the first and third carbon position.
Purine = pyramidine with imadazole ring fused to it.

Question 2

Difference between protein coding gene and protein non-coding genes?

Answer 2

Non-coding genes are transcribed but do not undergo translation - becoming rRNA, tRNA, microRNA…
Protein coding genes are transcribed then spliced to create mRNA purely composed of protein encoding genes - then moves out nucleus to cytosol where it undergoes translation in ribosomes.

Question 3

How do ribose and deoxyribose differ?

Answer 3

Ribose has OH attached to 2nd and 3rd carbons. Deoxyribose does not have OH attached to 2nd carbon, but has an H group.

Question 4

Precursors of DNA and RNA

Answer 4

RNA = ribonucleotide 5’ triphosphates ATP, CTP, GTP, UTP.
DNA = deoxyribonucleotide 5’ triphosphates dATP, dCTP, dTTP, dUTP

Question 5

What is RNA secondary structure and how does it happen?

Answer 5

When there are complementary bases within single stranded RNA and they pair with each other - intra strand base pairing.
E.g. stem loops, pseudo knots, hairpins.

Question 6

key aspects of DNA structure

Answer 6

1- hydrogen bonds
2- antiparallel
3- right handed helix
4- major groove and minor groove
5- roughly 10.5 bp per rotation
6- Double stranded, double helix
7- bases project into the middle
8- phosphate backbone

Question 7

Differences between DNA and RNA

Answer 7

Precursurs - uracil and thymine
Ribose and deoxyribose
Single + double stranded
RNA - intra base pairing
RNA - mRNA, rRNA, tRNA
DNA - genes

Question 8

How can DNA be denatured

Answer 8

• high pH, 0.1N NaOH
• Heat
• Enzymes

Question 9

DNA melting temperature

Answer 9

• Breaking double bond between AT - 2 degrees
  Breaking triple bond between CG - 4 degrees
  Almost all DNA strands are separated at 95 degrees - become single stranded.

Question 10

DNA annealing definition

Answer 10

When two single strands of complementary DNA are free in a solution, strands will bind to each other in their complementary areas as long as conditions are suitable.

Question 11

Explain how RNA polymerase synthesises RNA

Answer 11

1. RNA polymerase starts at promotor site and transcription starts at +1 site.
2. RNA polymerase either binds directly or indirectly to the promotor.
3. RNA polymerase recruits RNA precursors - NTPs.
4. RNA polymerase can synthesise RNA by using DNA as template
5. RNA polymerase synthesises RNA from 5’ to 3’ direction.
6. RNA polymerase catalyses the formation of 5’-3’ phosphodiester bonds between phosphate on 5’ carbon on NTP and 3’ OH on another nucleotide and released pyrophosphate.

Question 12

Function of DNA topoisomerase?

Answer 12

Relieves supercoiling of DNA in front of RNA polymerase, and restores proper level of coiling to DNA behind RNA polymerase.

Question 13

Explain how gene is transcribed into mRNA

Answer 13

• RNA polymerase initiates transcription by producing RNA
• Introns are removed by RNA splicing, and exons are rejoined in various ways to make different versions of mRNA to code for different versions of the protein.
• Strand is cleaved at polyA adenylation site on 3’ end (AAUAAA)
• Enzyme PolyA polymerase adds hundreds of adenines (polyA tail)
• 7 methyl guanine is added to 5’ end via 5’ to 5’ triphosphate bond –> mRNA is capped

Question 14

What is the +1 site

Answer 14

Position on DNA where RNA polymerase begins to synthesise RNA

Question 15

Terminator site in prokaryotes

Answer 15

Prokaryotes RNA forms a specific shape causing the RNA polymerase to dissociate from the DNA

Question 16

Transcription bubble

Answer 16

When RNA polymerase binds to promotor, DNA melts forming a transcription bubble. 14bp wide.

Question 17

Function of CAP

Answer 17

Translation initiation, protection, transport, splicing

Question 18

Function of polyA tail

Answer 18

Transport, protection + termination.

Question 19

Difference between mRNA in euk and prok?

Answer 19

Euk - splicing, cap, pokyA, not coupled.
Prok - Ribosome binding site, coupled.

Question 20

Describe the structure of the nucleosome

Answer 20

Histone octamer + DNA surrounding it.

Question 21

Describe the structure of histones

Answer 21

Histones are proteins that DNA is wound around.
Histone octamers are H3, H4, H2A, H2B.
H3 forms dimer with H4
H2A forms dimer with H2B.
H1 is not part of nucleosome or histone octamer, it is a linker protein that condenses nucleosomes and draws them together.

Question 22

DNA synthesis

Answer 22

• DNA polymerase uses one DNA strand as template to synthesise complementary strand.
• DNA polymerase cannot synthesise DNA from scratch - can only synthesise DNA onto 3’ end of an existing pieces of DNA or RNA called a primer.
• DNA polymerase can only attach an incoming dNTP onto 3’OH group of deoxyribose, thus DNA polymerase synthesises DNA in 5’-3’ direction.
• DNA polymerase also catalyses formation of phosphodiester bond between phosphate on 5’ carbon of incoming dNTP and 3’OH on primer, generating 5’-3’ phosphodiester bond.
• High energy dNTP is consumed, pyrophosphate is released.

Question 23

Proteins that work to repair mismatches

Answer 23

• MLH1, MSH2, MSH3, MSH6, PSM1, PMS2.
  These proteins excise mismatch and re synthesise DNA.

Question 24

Initiation of DNA synthesis

Answer 24

• Specific proteins bind to origin or replication. In eukaryotes, protein complex called ORC (origin recognition complex) binds to ori.
• Proteins at ori recruit additional proteins and melt DNA at ori.
• Helicases unwind DNA and form a replication bubble at ori. Single stranded binding protein binds to single stranded DNA and keeps it single stranded to enzymes can come in and work on it to replicate it.
• Proteins at ori recruit proteins that will replicate DNA
• RNA primer is synthesised at ori by DNA primase. When DNA is opened up, supercoiling builds up in areas flanking the opening. These needs to be relieved by Topoisomerase I and II (DNA gyrase in prokaryotes)
• After primer is synthesises, DNA polymerase can start DNA synthesis in 5’-3’
  direction.

Question 25

Leading strand:

Answer 25

newly synthesised DNA from RNA primer by DNA polymerase that continues all the way to the end of the DNA molecules

Question 26

Lagging strand:

Answer 26

. Okasaki fragments are started from RNA primer laid down by . . DNA primase.
. DNA primase synthesises primers at regular frequent intervals on the strand.
. DNA polymerases then synthesise new DNA from these primers in a 5’-3’ direaction and remove RNA primers.
. Eventually lots of small bits of DNA are synthesised adjacent to each other.
. DNA ligase seals adjacent DNA segments together.

Question 27

Key elements of chromosomes

Answer 27

• 2 telomeres
  
  • A centromere
  • One or more origins of replication

Question 28

Telomere + its function

Answer 28

large nucleoprotein complex at the ends of eukaryotic chromosomes.
- Stable
- Protect chromosome from damage & degradation
- Prevents end of chromosome fusing with other chromosomes.
- Made of TTAGGG repeated thousands of times

Question 29

How does telomerase replicate end of chromosomes

Answer 29

1. Telomerase binds to end of chromosome at 3’ end. DNA is being made on RNA template -> reverse transcriptase
2. Telomerase uses a sequence of its own RNA to extend 3’ end of chromosome by TTAGGG
3. Telomerase translocates 6 bases
4. Repeats this process

Question 30

Retroviruses

Answer 30

Copy their RNA into DNA.
- Have positive RNA
- Copied into DNA by reverse transcriptase
- DNA inserted into genome of host, and inserted genome is transcribed into +RNA by host RNA polymerase.

Question 31

Why is the mutation rate in RNA viruses very high?

Answer 31

RNA polymerase does not have proofreading ability, mismatch repair cannot be carried out on RNA genome.

Question 32

Double stranded RNA viruses:

Answer 32

Viruses that copy their RNA into new RNA
Positive strand RNA viruses: RNA can be directly translated into protein
Negative strand RNA viruses: RNA must first be converted into positive RNA for translation.

Question 33

Type of mutations in virus RNA

Answer 33

Missense mutation - changes codons. Varied effects, can be positive or negative.
Silent mutation - has no effects
Nonsense mutation - changes codon to stop codon. Typically stops protein from working.

Question 34

Nucleotide triphosphate inhibitors

Answer 34

Molecules that resemble those used to make dNTPs/ NTPs but inhibit their production.

Question 35

Antimetabolites

Answer 35

Prevent synthesis/ formation of pyrimidine and purine.

Question 36

Inhibitors of chain elongation

Answer 36

Incorporated into DNA by DNA polymerase but cause chain termination as incoming nucleotides cannot be added onto them.

Question 37

Genome

Answer 37

DNA sequence, the same in all somatic cells besides low levels of mutation accumulation

Question 38

Methylome

Answer 38

Methylation pattern. Different in cells of different tissues. Changes due to signalling factors, environment, age …

Question 39

Proteome

Answer 39

Proteins expressed, differs in cells from different tissues. Also changes in cells due to different factors like signalling, environment etc…

Question 40

Transcriptome

Answer 40

RNA transcribed from all genes in a cell or tissue. Differs between cells of different tissues.

Question 41

Epigenome

Answer 41

Contains
- DNA
- nucleosome
- Pattern of all methylated cytosines
- Pattern of all modifications to chromatin
- Transcription factors bound to chromatin
- Remodelling complexes bound to chromatin.