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The human genome > Genome chemistry, epigenetic & DNA methylation (meg) > Flashcards

Genome chemistry, epigenetic & DNA methylation (meg) Flashcards

1
Q

What can genomes be made of?

A

DNA or RNA

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2
Q

Name 5 viruses made up of single stranded RNA

A

HIV
Flu
Ebola
Polio
Measles

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3
Q

Name 2 viruses that are made up of double stranded RNA

A

blue tongue
rotavirus

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4
Q

Name 3 viruses that are made up of single stranded DNA

A

M13
ɸX174
sea water viruses

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5
Q

Name 4 viruses that are made up of double stranded DNA

A

Complex viruses:
- Herpes
- Smallpox
- bacteriophage T4
- lambda phage

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6
Q

What are the genomes of cellular made of?

A

Double stranded DNA

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7
Q
A
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8
Q

What is the corona virus genome made of?

A

Single stranded RNA

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9
Q

What are the 2 levels of nucleic acid chemistry?

A
  1. Individual bases = nucleotides
  2. Higher order structure (double helix)
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10
Q

What is this picture showing?

A

RNA nucleotide (need to be able to draw)

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11
Q

What defines an RNA nucleotide?

A

the hydroxyl group at the 2’ position on the ribose sugar

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11
Q
A
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12
Q

How many carbons are there in ribose / deoxyribose sugars?

A

5

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13
Q

What is the difference between an deoxyribose sugars and ribose sugars?

A

Deoxyribose = oxygen is missing from 2’ prime position so just a single hydrogen = resulting in important consequences and making DNA much more stable than RNA

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14
Q

What makes an deoxyribose sugar more stable than a ribose sugar?

A

hydroxyl group on RNA can catalyse the hydrolysis of the phosphodiester bond

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15
Q

Name all the pyrimidine bases?

A

C
T
U

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16
Q

Outline the structure of pyrimidine bases

A

2 nitrogens and 4 carbons in pyrimidine labelled counter clockwise

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17
Q

Name the 2 pyrimidine bases in DNA

A

Cytosine
Thymine

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18
Q

Outline the structure of cytosine and thymine

A

Cytosine = carbonyl group (C=O) at position 2’, at 4’ position = amino group

thymine = same carbonyl group at 2 position + carbonyl group at 4’ position and at 5’ position = methyl group

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19
Q

In terms of bases, what is the difference between RNA and DNA?

A

In RNA, it is uracil instead of thymine (DNA)- both pair with adenine

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20
Q

Describe what was discovered (2023 nobel peace prize in medicine)

A

In RNA instead of uracil- many bases are modified to have pseudo-uracil

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21
Q

What is pseudo-uracil?

A

looks like uracil but instead of having base attached to ribose sugar through nitrogen, it is attached through carbon

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22
Q

How is the base attached to ribose sugar through carbon in pseudo-uracil?

A

Enzyme enables this- rotates the base and re-attaches through the carbon atom

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23
Q

Why was this discovery important?

A
  • mRNA with pseudo uracil in it translated more efficiently than non-modified mRNA
  • cells don’t promote an inflammatory response- way of discriminating between viral DNA which is unmodified and cellular DNA which is modified
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24
Q

Why did this discovery win the 2023 Nobel Peace Prize?

A

all RNA COVID vaccines used pseudo-uracil instead of uracil- it worked because without pseudo uracil much less protein would have been produced and the cells would have promoted an inflammatory response

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25
Q

Name the 2 main differences between RNA and DNA

A
  1. DNA has thymidine (thymine) - RNA has uridine (uracil)
  2. 2’ position of ribose sugar in DNA has a hydrogen atom while RNA has a hydroxyl group
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26
Q

Describe the structure of Deoxythymidine triphosphate

A
  • 3 phosphate groups
  • Thymidine- 5’ position has methyl group and carbonyl groups at 2’ and 4’ positions
  • No hydroxyl group on sugar = DNA nucleotide
  • Carbon atom present at 2’ position on deoxyribose sugar
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27
Q

What are the 3 key features of nucleic acids?

A
  • Polarity = 5’ end distinct from 3’ end
  • Bases are linked by phosphodiester bonds = 2 ester linkages
  • Glycosidic bond- bond between 1’ carbon on ribose sugar and 1’ position nitrogen on base
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28
Q

What is the direction of a nucleic acid sequence?

A

5’ to 3’ direction

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29
Q

What is between bases in double stranded nucleic acids?

A

hydrogen bonds

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30
Q

How many bonds are between each type of base pair?

A

T + A = 2 hydrogen bonds
C + G = 3 hydrogen bonds

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31
Q

Describe the arrangement of double stranded DNA

A

2 DNA strands have an antiparallel arrangement (one strand 5’ to 3’ and other strand is 3’ to 5’) and are wrapped around each other in plectonemic coil

32
Q

Why is DNA a good molecule to encode genetic information and make stable genomes?

A
  1. DNA is more stable than RNA because of the presence of 2’ OH group in ribose sugar in RNA makes RNA alkali labile (= it breaks up)- 2’ OH acts as a catalyst for hydrolysis
    - Hydroxyl group in presence of alkali could cause nucleophilic attack e.g. on a phosphate and in doing so, it would displace an oxygen next to the phosphate group- could result in phosphodiester backbone being cleaved and in turn impact the condition of the oligonucleotide
  2. DNA has thymine instead of uracil- cytidine is unstable and in the presence of thymine allows the products of cytidine instability to be identified and removed = repaired
33
Q

Define: cytidine

A

= nucleotide containing cytosine

34
Q

what is an oligonucleotide?

A

= short chain of nucleotides

35
Q

what is cytosine deamination?

A

hydrolytic reaction = converts cytosine into uracil as amino acid is converted into a carbon r group

36
Q

Is cytidine deamination a physiological process or a non-physiological process?

A

non-physiological process

37
Q

What is the difference between physiological and non-physiological processes?

A

physiological = characteristic of or inherent to the normal functioning of living organisms

38
Q

Explain how cytosine deamination is different in DNA and RNA

A

Difference lies in the outcomes of the process:
RNA = end up with uracil

In DNA- have thymine instead of uracil, but when cytosine deamination happens, we get uracil. The cell immediately knows that uracil should not be in DNA due to presence of thymine:
- The uracil is identified and repaired back into thymine
- So presence of thymine in DNA to enable cell to recognise the product (=uracil) of cytosine deamination

39
Q

Name the enzyme that removes uracil in DNA and how does it do this?

A

uracil glycosylase- breaks glycosidic bond between sugar and the uracil = 1st step of base excision repair = BER

40
Q

What does base excision repair (BER) also do?

A

Removes oxidised and alkylated bases

41
Q

Describe the process of BER

A

BER removes the U and restores C:
1. By cutting the glycosidic bond = get apurinic/apyridinic site

  1. Enzymatic machinery comes along and removes apurinic/apyridinic site
  2. Repairs it and puts in a thymine- 2 types of repair reaction:
  • Long patch
  • Short patch
42
Q

What came before the other- RNA or DNA?

A

Good evidence to suggest that RNA came before DNA

43
Q

Outline the evidence to suggest that RNA came before DNA

A
  • both can assemble on the basis of complementarity + store info but only RNA is significantly catalytically active
  • all descended from RNA molecules- RNA performs both catalysis as starting point for life + in lab you can evolve RNA molecules = can make self-replicating RNA molecules
  • DNA precursors are made from RNA precursors by ribonucleotide reductase
  • 2 different thymidylate synthetases that methylate dUMP in many organisms suggesting T evolved twice
  • PBS1 and PBS2 phage of B.subtilis contain U instead of T in their DNA- suggests its an ancestor for DNA
44
Q

What does ribonucleotide reductase do?

A

Reduces ribonucleotides

45
Q

Name some more reasons why double-stranded DNA is a good choice to encode genetic information

A
  • Double stranded DNA does not fall apart when phosphodiester backbone nicked since it is stabilized by hydrogen bonds and by hydrophobic interactions between the faces of the individual base pairs
  • Nucleotide bases are protected from aqueous phase by phosphodiester backbone- in DNA = potentially reactive groups attached to bases e.g. amino groups, carbonyl groups but in double helix these are protected by phosphodiester backbone unlike single-stranded nucleic acids
  • Double helix can be melted under physiological conditions- if it didn’t melt then genetic info couldn’t be accessed and transcribed- perfect balance between stability and instability
46
Q

Name the 3 different double helices that double stranded nucleic acids can form

A

A DNA
B DNA
Z DNA

47
Q

What is the main form of double stranded DNA helix?

A

B-DNA

48
Q

Describe the structure of B DNA and why is this beneficial?

A

Have a minor groove (can’t see the bases) and a major groove (can see bases)- good because transcription factors can read base sequence
Do this by binding through the major groove

49
Q

How is Z DNA formed?

A

alternating purines and pyrimidines

50
Q

What 2 types of interaction stablises the double helix?

A

Both non-covalent interactions:
1. Hydrogen bonds between bases
2. Hydrophobic stacking interaction between faces of the bases

51
Q

Why are hydrophobic stacking interaction between faces of the bases stronger at high salt concentrations?

A

stronger at high salt concentration

52
Q

if temperature is raised what will happen to double stranded DNA?

A

Double stranded DNA will melt into 2 strands if temp is raised

53
Q

After melting double stranded DNA can this be reversed?

A

reaction reversible = hybridisation = 2 double strands will bond back together

54
Q

Why is nucleic acid hybridisation fundamental in biology?

A

important in mRNA translation, tRNA transcription- codon (mRNA) and anticodon (tRNA) interactions, homologous recombination = hybridisation of single strands of DNA

55
Q

Why is nucleic acid hybridisation fundamental in molecular genetics?

A
  • PCR- depends upon hybridisation of the primers,
  • Southern blotting- depends upon hybridisation,
  • FISH- depends upon hybridisation of single stranded probe with denatured single stranded chromosomes,
  • DNA sequencing- primers are annealed to single stranded DNA (same as PCR)
56
Q

What is this picture showing?

A

Absorption at 260nm increases as double stranded DNA melts = hyperchromicity

57
Q

What is DNA melting temperature sensitive to?

A

GC content

58
Q

Why is melting temperture sensitive to GC content?

A

GC rich DNA will melt much more at a higher temperature than AT rich DNA- because there are more hydrogen bonds between guanine and cytosine base pairs

59
Q

is cytidine deamination mutagenic?

A

Yes but is sometimes exploited biologically in physiological processes i.e Cytidine deamination causes mutations but the consequences of mutational processes are part of normal physiology of organisms

60
Q

What group of enzymes catalyse cytidine deamination?

A

cytidine deaminases

61
Q

Explain what cytidine deaminases are involved in

A
  1. RNA editing- important in birds also
  2. Innate immunity to retroviruses and transposable elements- so our cells have systems for protecting themselves
  3. Adaptive immunity = production of antibodies and antigen specific T cells- cytidine deaminases are used by adaptive immune system as part of normal process immunoglobulin gene diversification (=antibody diversification)
  4. Cancer- cytidine deaminases sometimes get mis expressed = mutations that give rise to cancer
62
Q

How were cytidine deaminases discovered?

A

in RNA editing

63
Q

Name the 2 forms of the apolipoprotein B (ApoB)

A

ApoB100
ApoB48

64
Q

Where is ApoB100 synthesised?

A

Liver

65
Q

Name the 2 domains on ApoB100

A

lipoprotein assembly
LDL receptor

66
Q

What is the ApoB100 responsible for?

A

transporting lipids from liver to muscle tissues for production of energy

67
Q

How do ApoB100 carry out their function?

A

LDL receptor domain on ApoB100 is bound to LDL receptor on tissue (particularly on muscle)- it enables muscle to take up the lipid on VLDL synthesised by the liver

68
Q

Where is ApoB48 synthesised?

A

Small intestine

69
Q

does ApoB48 have LDL receptor domains?

A

48% of protein lacked LDL receptor domain

70
Q

Name the domains in ApoB48

A

lipoprotein assembly domain only

71
Q

What is the ApoB48 responsible for?

A

carrying lipids from the gut back to liver

required for synthesis of chylomicrons by gut

72
Q

Define LDL and VDL

A

LDL = low density lipoprotein
VLDL = very low density lipoprotein

73
Q

What was the explanation for the 2 forms of ApoB?

A

The full length message encoded the Apo-B100 with the LDL receptor binding domain- it included all of the sequence corresponding to the exons in the gene

Then mRNA encoding the Apo-B48- had same length but one residue of cytidine in mRNA sequence which had been deaminated to uracil

  • Result of deamination + presence of uracil = CAA codon (= glutamine) was converted into a stop codon by APOBEC-1- so abbreviated that position giving rise to Apo-B48
74
Q

What is AID?

A

= activation induced deaminases (protein)

75
Q

What are AID important in?

A

immunoglobulin gene hypermutation

76
Q

Describe immunoglobulin gene hypermutation

A

1st produce antibodies they have low affinity but in mature B cells IG hypermutation takes place and the genes encoding the antibodies get further mutated- as a result of this = antibody response matures and become progressively of higher affinity

77
Q
A

The human genome (20 decks)

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