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BB10006 Cell and Molecular Biology > Transcription and translation > Flashcards

Transcription and translation Flashcards

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1
Q

DNA requirements to be genetic material

A

-Must carry information
-Must replicate
-must allow info to change-mutation
-must govern the expression of the phenotype-gene function
Stores information in sequence of bases

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2
Q

What does LDL cause?

A

If people have this single base pair change, change the sequence of amino acids asn to serine

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3
Q

What is serine associated with?

A
  • High cholesterol
  • high LDLS
  • coronary heart disease
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4
Q

What is an example of personalised medicine?

A

CYP 2D6 gene sequence

  • amino acid changed to stop codon, enzyme cannot break down certain drugs
  • doctors then can prescribe a different tailored drug for patient
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5
Q

In replication what keeps DNA from annealing?

A

-Single-Stranded DNA proteins

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6
Q

Who discovered the mechanism in biological synthesis of DNA replication?

A

Kornberg in 1959

He discovered the process of complimentary base pairing and how it could happen

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7
Q

What is the mechanism of DNA replication?

A
  • Initiation-proteins bind to DNA and open up helix, prepare DNA for complimentary base pairing.
  • Elongation-proteins via DNA polymerase connect sequences of nucleotides into continuous new strand of DNA.
  • Termination-proteins released the replication complex.
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8
Q

What are the rules of DNA Replication?

A 
-Semiconservative
– starts at DNA origin
– can be uni or bidirectional
-semi discontinuous
– always 5’-3’ direction
-RNA primers required
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9
Q

What is semiconservative replication and who discovered it?

A

Meselson -Stahl Proved half of DNA comes from each parent

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10
Q

Where does DNA replication start from?

A

Starts at an ORI sequence
- Initiator proteins identify specific base sequences of DNA called sites of origin
– highly conserved
-prokaryotes have single ori sites eukaryotes have multiple ori sites

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11
Q

What is the difference between replication in bacteria and humans?

A
  • In bacteria much faster application
  • enzymes not as good as proofreading
  • replication in humans quicker if multiple firing of ori sites
  • more accurate in humans
  • initiation proteins bind to initiator sites
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12
Q

What is semi discontinuous replication?

A
  • Antiparallel strands replicated simultaneously
  • leading strand synthesis continuously in 5‘-3‘ direction
  • lagging strand synthesis in fragments in 5’-3’ direction
  • Lagging strand is slower, moving in opposite direction to fork, synthesis done in OkaZaki fragments
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13
Q

What is an Okazaki fragment?

A

-Short sequences of DNA nucleotides which are synthesised discontinuously later linked by DNA ligase to create lagging strand

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14
Q

Why does DNA replication only occur in five prime to 3 prime direction?

A
  • Breakdown of phosphate provides energy
  • must break triphosphate down for replication to work - only can occur in 5’-3’ direction
  • If it was 3’-5’ enzyme would get added but subsequent dNTPs is cannot bind, they are blocked
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15
Q

What is base mismatch?

A
  • Polymerase has multiple subunits involved in proofreading that right bases are added
  • it will then remove mismatched bases
  • this is why replication is slower in Eukaryotes
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16
Q

Why are RNA Primers required for DNA synthesis?

A
  • RNA primer required, lay down for DNA replication to occur
  • done by a primase
  • One primer on leading strand, loads of primer is on lagging as every time DNA is added needs a primer
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17
Q

Function of topoisomerases

A

Prevents torsion by DNA breaks

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18
Q

Function of helicase

A

Separates two strands

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19
Q

Function of primase

A

RNA primer synthesis

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20
Q

Function of single-stranded binding protein

A

Prevents re-annealing of single strands

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21
Q

Function of DNA polymerase

A

Synthesis of new strand

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22
Q

Function of tethering protein

A

Stabilises DNA polymerase

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23
Q

Function of DNA ligase

A

Phosphodiester bond linkage

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24
Q

In prokaryotes the genes are usually…

A

Switched on – codes for components for production of tryptophan
-gene is transcribed in operons

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25
Q

In eukaryotes genes are usually…

A
  • Switched off

- not all genes code for proteins

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26
Q

Where does transcription occur in prokaryotes? And what occurs? Eg. Tryptophan

A

Cytoplasm due to lack of nucleus

Tryptophan -

  • Transcription starts at one point and transcript is made from operon
  • Entire transcript made in one go in operon
  • genes are switched on
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27
Q

Features of transcription in eukaryotes

A
  • Linear-each gene responsible for tryptophan on different chromosomes
  • gene is usually switched off
  • series of mRNA transcripts which are then translated into proteins
  • making of mRNA transcript occurs in nucleus –Primary transcript
  • mRNA-out of nucleus for translation
  • not all genes code for protein
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28
Q

What does micro RNA do?

A

Bonds to another RNA molecule and destroys it so that no protein is made

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29
Q

Where does the untranslated regions lie?

A

In between initiation codon

- its not translated

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30
Q

What are the regulatory regions in transcription?

A
  • Tata box
  • GC box
  • Order in which boxes placed tell you this is regulatory region
  • highly conserved regions dictate whether transcription should occur
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31
Q

What are the coding regions of DNA in transcription?

A
  • AG and GT at beginning and end of exon, the bits inbetween are introns
  • All species have methionine at start (ATG)
  • At end highly conserved stop codons
  • base sequence at end is AATAA
32
Q

What are the three stages in the mechanism of transcription?

A

Initiation
Polymerisation
Termination

33
Q

What occurs in initiation in transcription?

A

RNA polymerase binds to promoter Along with transcription factors and opens helix

34
Q

What occurs in polymerisation in transcription?

A

De novo synthesis using rNTPs as substrate

- chain elongation in 5’-3’ direction

35
Q

What occurs in termination in transcription?

A

Stops at termination signal, stop codons

-one stand always a template

36
Q

What occurs in maturation of primary mRNA transcript?

A
  • RNA capping
  • polyadenylation
  • splicing

-end up with mRNA transcription with cap, polyadenine tail and no introns

37
Q

What is capping?

A
  • Addition of 7-methyl-guanosine at 5’ end

- Mediated by guanylyltransferase

38
Q

What does capping do?

A

-mRNA highly prone to degradation, mRNA highly unstable compared to DNA
-In protects against the degradation, serves as recognition site for ribosomes, transport mRNA out.
– MRNA goes via nuclear pore into cytoplasm using enzymes

39
Q

What is tailing?

A
  • Addition of Poly(A) residues at 3’ end

- Driven from energy from ATP

40
Q

How is the poly a tail attached?

A
  • Poly(A) polymerase, cleavage and polyadenylation specificity factor attaches Poly(A) generated from ATP
  • Add series of A’s to 3’ end
41
Q

What is splicing?

A

-Removal of introns by snRNPs and spliceosomes

42
Q

How does splicing occur?

A
  • One cut made, loop made from free-floating end, second cut from route of Lariat to splice intron out
  • recognise Exon intro boundaries
  • Once spliced you have mature RNA
43
Q

What does alternative splicing allow?

A

Multiple transcripts from a single gene

44
Q

What are housekeeping genes?

A

-Genes which are made all the time e.g. tubulin

45
Q

What three things does translation require?

A

1) mRNA
2) aminoacyl-transfer RNA (aatRNA)
3) Ribosomes

46
Q

In translation what do ribosomes form?

A

A catalytic domain into which tRNAs enter with the attached amino acids

47
Q

What does degenerate mean?

A

More than one triplet per amino acid

48
Q

Features of tRNA molecule

A
  • One tRNA per amino acid
  • tRNA must be charged
  • D loop, T loop, has amino acid tagged onto three prime end
  • Two active sites, one for the amino acid one for tRNA
49
Q

What must be done to tRNA?

A

1) Covalently attached correct amino acid to tRNA specified by anti codon
2) select the correct charged tRNA specified by mRNA
- coupling of correct amino acid done via aminoacyl tRNA synthetases

50
Q

When selecting the correct amino acid (done via aminoacyl tRNA Synthetases) what doesn’t the 3rd base have to do?

A

Conform to Watson Crick base pairing

51
Q

What occurs in initiation in translation?

STAGE 1 - FORMATION OF 43S pre-initiation complex

A

-mRNA sequence signals where protein synthesis begins

  • 40s-eIF3 bound by eIFA to ternary complex of tRNA (methionine, eIF2 and GTP)
    -First amino acid always methionine, energy from GTP not ATP
    – Cofactor 2 bound to GTP
    -Anti codon loop recognises its complement
52
Q

What occurs in initiation in translation?

STAGE 2 - FORMATION OF initiation complex

A
  • cap Binding of mRNA to 43S
  • mRNA has cap and tail, have protein is bound to them called PABP, bind on 3’ end of sequence to A’s
  • Initiation factor complex comes in (eIF4) and binds to 5’ end of cap
  • Within complex head and tail link to form a loop
  • allows for charged methionine to bind
53
Q

What occurs in initiation in translation?

STAGE 3 - Positioning at start codon

A
  • Initiation complex unwinds mRNA using eIF4 helicase
  • Initiation complex stops at start site AUG
  • Recognition allows an irreversible GTP hydrolysis of eIF2 preventing further unwinding
  • All secondary structures removed, Helicase to prime apart
54
Q

What is the highly conserved sequence to allow the complex to know the true AUG start codon?

A

ACCAUGG KOZAK SEQ

55
Q

What occurs in chain elongation in translation?

STAGE 1 - aatRNA binds

A
  • aatRNA binds to A site on ribosome by base pairing with codon
  • Methionine start codon
  • EF1 driven by GTP energy currency, bring correct charged tRNA ahead of the methionine
  • 1st initiation methionine called met i
56
Q

What does EF stand for?

A

Elongation factor

57
Q

What occurs in chain elongation in translation?

STAGE 2 - Conformation change

A
  • Confirmation change in the ribosome, induced by GTP hydrolysis of eF1a
  • In order for amino acids to form peptide bond with the next amino acid’s ribosome changes shape
  • EF1 Dissociates as its job is done
58
Q

What occurs in chain elongation in translation?

STAGE 3 - Transpeptidation

A
  • C Terminal of polypeptide uncoupled from P site tRNA and peptide bond transferred to amino acid on A site tRNA Catalysed by Peptidyltransferase
  • No charged tRNA here
59
Q

What occurs in chain elongation in translation?

STAGE 4 - Translocation

A
  • GTP Hydrolysis of EF2 to cause second confirmational change
  • P site tRNA transferred to E site
  • Simultaneous transfer of A site tRNA move to P site
  • shape changeX EF2 driven by GTP
60
Q

What occurs in step three of translation - termination?

A
  • Release factors recognise and bind to stop codons
  • induces peptidyl transferase to transfer peptidyl group to water instead of aatRNA
  • Uncharged tRNA released from ribosomes
  • Inactive ribosome then release mRNA
61
Q

Where are proteins in translation made?

A

Polyribosomes

62
Q

Why does mRNA have a loop confirmation in translation?

A

Translation increased efficiency

63
Q

Post-translational modifications-protein folding

A
  • Nascent protein folded

- Mediated by HSP 70/HSP 60 complexes

64
Q

Post-translational modifications – covalent modifications

A

-Addition of side groups
-Kinases add phosphate group to certain target proteins
– sidechains added on
– dictate activity/lifespan of proteins

65
Q

Post-translational modifications - proteolytic cleavage

A

-Activate some in active precursors

66
Q

In proteolytic cleavage what do caspases do?

A

Trigger apoptosis

67
Q

In proteolytic cleavage what do zymogens do?

A

Inactive precursors, only activated when cleaved

68
Q

What is the a site in a ribosome?

A

acceptor site – binds to aminoacyl tRNA which holds new amino acid to be added

69
Q

What is the P site in a ribosome?

A

Peptidyl site-binds to tRNA holding growing polypeptide of amino acid

70
Q

What is the e site in ribosome?

A

Exit site-threshold, final transitory step before tRNA now bereft of amino acid is let go by ribosome

71
Q

Antibiotics that inhibit translation only found in prokaryotes

A

Streptomycin-prevents initiation elongation

Chloramphenicol-Blocks peptidyl transferase in mitochondria

72
Q

Antibiotics that inhibit translation only found in eukaryotes

A

Cycloheximide-Blocks translocation only in cytosol

73
Q

Antibiotics that inhibit translation found in both prokaryotes and eukaryotes

A

Puromycin – causes premature release of polypeptide

74
Q

Which of the following scientists first discovered that the percentage of GC is constant in any given species.

A

Erwin Chargaff

75
Q

Small nucleolar RNAs are involved in

A

rRNA modification and processing

76
Q

The ability of DNA to denature is important for

A

DNA synthesis
Nucleic acid hybridization experiments
RNA synthesis
Polymerase Chain Reaction

77
Q

What occurs in initiation in translation?

STAGE 4 - Association of large subunit (60S)

A

Irreversible GTP hydrolysis mediates the association of 60S-eIF6 (large subunit ) to the small subunit by the action of eIF5
This becomes the P site

BB10006 Cell and Molecular Biology (5 decks)

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