Biochemistry of Nucleic Acids

Question 1

what is the central dogma?

Answer 1

DNA > RNA > Protein

Question 2

difference between ribose and deoxyribose?

Answer 2

ribose has 2 OH groups, deoxyribose only has 1

Question 3

how are carbons numbered in nucleotides/nucleosides?

Answer 3

from amino end to phosphate end

Question 4

DNA building blocks vs RNA building blocks?

Answer 4

DNA = dATP, dCTP, dGTP, dTTP
RNA = ATP, CTP, GTP. UTP

Question 5

how does polymerisation occur in DNA?

Answer 5

phosphodiester bond is formed between a free 3’ OH group on the above nucleotide and a 5’ triphosphate on ATP, leaving 2 diphosphate
consumes 2 high energy bonds

Question 6

give an example of a nucleotide analogue used as a drug and describe how it works

Answer 6

ZDV/AZT/Retrovir (analogue of thymidine)

incorporated into growing viral DNA but lacks 3’ OH group so chain elongation is terminated

Question 7

how many bond between A/T and C/G?

Answer 7

A-T = double bond
C-G = triple bond

Question 8

replication is conservative, true or false?

Answer 8

false

only semi conservative

Question 9

what catalyses DNA replication?

Answer 9

DNA polymerase

can only add to existing nucleic acid and require an RNA primer to start replication

Question 10

in which direction is DNA replicated?

Answer 10

bidirectional

always 5’ to 3’

Question 11

building blocks of DNA replication?

Answer 11

dATP, dTTP, dCTP, dGTP

Question 12

what synthesises an RNA primer?

Answer 12

primase

Question 13

what is the exonuclease activity of DNA polymerase?

Answer 13

moves 3’ to 5’ removing incorrect nucleotides

improves error rate

Question 14

most abundant RNA?

Answer 14

rRNA (80%)
forms ribosomes
tRNA = (15%)
mRNA = (5%)

Question 15

describe structure and function of tRNA

Answer 15

clover leaf structure when flattened

specific amino acid is attached to 3’ end (dependant on anti-codon sequence)

Question 16

what are the 3 types of eukaryotic RNA and how can they be distinguished?

Answer 16

Pol I, ii and iii (Pol ii makes all mRNA)

by sensitivity to toxins like alpha amanitin

Question 17

what is TBP?

Answer 17

TATA box Binding Protein
recognises TATA box (present in promotor region)
part of TFIID
introduces kink into DNA providing landing platform for further transcription factors and RNA polymerase

Question 18

what is TFIID?

Answer 18

general transcription factor required for all Pol ii transcribed genes

Question 19

how is transcription initiated?

Answer 19

needs additional general transcription factors
Pol ii and TFIIF extend transcript on their own
TFIID remains at promotor, a new initiation complex can now assemble
allows transcription at low, basal rates

Question 20

how is the transcribed strand elongated?

Answer 20

transcription bubble moves along DNA from 5’ to 3’ (DNA unwound in front of polymerase and rewound behind it)

Question 21

how is transcription terminated?

Answer 21

new RNA makes a stem loop structure (followed by stretch of U’s)
specific enzyme cleaves the finished RNA
RNA released and polymerase dissociates

Question 22

how is transcription regulated?

Answer 22

needs specific transcription factors (DNA binding proteins with DNA binding domain and transcriptional activation domain)
bind to specific DNA sequences in vicinity of promoter (enancers)

Question 23

describe coordinated gene expression

Answer 23

a stress (e.g hormone stimuli, cellular stress etc) activates transcription of a regulatory protein through a stress sensitive transcription factor
binding of the regulatory protein to the stress regulatory element (SRE) stimulates transcription of genes A, B and C
Genes A, B and C produce different proteins which participate in the stress response

Question 24

give 2 examples of coordinated gene expression

Answer 24

steroid receptors (on binding, ligand (steroid) moves to nucleus and binds to DNA at steroid response elements (SRE)
glucocorticoid receptors (free steroids enter target cells via diffusion, bind to inactive steroid receptor in cytoplasm, activates receptor, translocates to nucleus, binds to response elements as homodimer, coordinated regulation of set of genes)

Question 25

when does splicing take place?

Answer 25

after transcription, before translation

Question 26

what happens at the 5' and 3' ends of mRNA?

Answer 26

Poly(A) tail (AAAAA) added at 3' end | modified GTP "cap" added at 5' end

Question 27

what is the difference between transcription/translation in prokaryotes and eukaryotes?

Answer 27

compartmentalised in eukaryotes - transcription in nucleus - translation in cytosol occurs anywhere in prokaryotes

Question 28

degenerate vs unambiguous?

Answer 28

``` degenerate = amino acid has more than one codon unambiguous = each codon has only one amino acid (or a stop) ```

Question 29

start vs stop codons?

Answer 29

``` start = AUG stop = UAA, UAG, UGA ```

Question 30

what are the 3 different reading frames of RNA (e.g if there are an extra 2 bases which cant complete a codon)?

Answer 30

2 bases at the right end 1 base either side 2 bases at the left end

Question 31

what are the components of translation?

Answer 31

``` amino acids tRNAs Aminoacyl-tRNA synthetases Specific protein factors for - initiation of protein synthesis - elongation - termination ATP and GTP (energy) Ribosomes mRNA ```

Question 32

what does aminoacyl tRNA synthetase do?

Answer 32

attaches amino acid to its tRNA via covalent bond | at least 1 per amino acid

Question 33

how many rRNA molecules are contained within each ribosome?

Answer 33

4 | 3 in bacteria

Question 34

what are the components of a ribosome?

Answer 34

4 rRNA molecules | proteins

Question 35

which has bigger ribosomes, prokaryotes or eukaryotes?

Answer 35

eukaryotes

Question 36

3 tRNA binding sites of ribosomes?

Answer 36

Exit Peptidyl Aminoacyl

Question 37

initiation of translation?

Answer 37

needs initiation factors (IFs) energy from GTP hydrolysis small ribosomal subunit binds to 5' end of mRNA moves along mRNA until start codon met initiator tRNA with UAC anticodon base pairs with start (carries methionine) large subunit joins assembly and initiator tRNA located in P site

Question 38

elongation of translated protein?

Answer 38

``` elongation factor (EF-1alpha) brings next aminoacyl-tRNA to A site (anticodon-codon) GTP hydrolysed, EF released from tRNA second elongation factor (EF-1betagamma) regenerates EF1alpha ```

Question 39

what catalyses the peptide bond formation between amino acids and where does this occur?

Answer 39

peptidyl transferase | occurs in P and A sites

Question 40

how does the ribosome move along the mRNA?

Answer 40

elongation factor EF-2 moves it along by 1 triplet at a time

Question 41

what is the pathway a tRNA takes in a ribosome of a growing peptide?

Answer 41

moves from A to P and then the empty tRNA moves to the E site where it can exit and become reloaded with an amino acid

Question 42

where are the A, P and E sites located on a ribosome?

Answer 42

``` A = furthest towards 3' end E = furthest towards 5' end P = in the middle ```

Question 43

how does termination occur?

Answer 43

when A site encounters a stop codon (no aminoacyl tRNA pairs with a stop codon) release factor RF binds stop codon (GTP hydrolysis) finished protein cleaved off tRNA rRNA, mRNA and tRNA dissociate from each other

Question 44

what is a polysome?

Answer 44

mRNA with several ribosomes attached

Question 45

point mutation?

Answer 45

change in single DNA base

Question 46

missense mutation?

Answer 46

causes change in amino acid sequence | can change protein function (sickle cell anaemia)

Question 47

Nonsense mutation?

Answer 47

creates new termination codon | changes length of protein due to premature stop of translation

Question 48

silent mutation?

Answer 48

no change in amino acid sequence due to degeneracy of genetic code no effect on protein function

Question 49

frameshift mutation?

Answer 49

addition or deletion of single base (or 2) | changes reading frame of translation into protein

Question 50

types of chromosomal mutations? how are they different?

Answer 50

``` deletions duplications inversions translocations affect larger portions of genome ```

Question 51

what are the 3 options for a finished protein?

Answer 51

targeting (moving to final cellular destination, depends on amino acid sequence) modification (adding further functional groups) degradation (unwanted or damaged proteins have to be removed)

Question 52

what do free ribosomes do and where are they found?

Answer 52

``` make proteins for - cytosol - nucleus - mitochondria - translocated post-translationally found in cytosol ```

Question 53

what do bound ribosomes do and where are they found?

Answer 53

``` make proteins for - plasma membrane - ER - Golgi apparatus - secretion - translocated co-translationally found on rough ER ```

Question 54

what are the 2 destinations for newly synthesised proteins?

Answer 54

to organelles/cytosol | to rough ER

Question 55

give 4 examples of post translational modifications that can occur in the ER

Answer 55

glycosylation (adding/processing carbohydrates in ER an Golgi) forming disulphide bonds folding/multiunit assembly proteolytic cleavage (in ER, Golgi and secretory vesicles)

Question 56

what results from misfolding of alpha1-antitrypsin in the ER?

Answer 56

hereditary emphysema

Question 57

give a clinical example of protein targeting

Answer 57

I-cell disease (recessive) proteins destined for lysosomes not properly sorted and end up secreted from the cell lysosomes cant digest material and become clogged death before age 8