Transcription & Translation

Question 1

RNA and DNA differences?

Answer 1

Thymine has an extra methyl group compared to uracil which makes it more stable. RNA is usually single stranded whereas DNA is double stranded. Ribose sugar in RNA, deoxyribose sugar in DNA.

Question 2

How do RNA polymerases copy genes?

Answer 2

By recognising promoters and terminators.

Question 3

What is a gene?

Answer 3

Basic unit of biological information. A specific segment of DNA that encodes an RNA or protein.

Question 4

What is a chromosome?

Answer 4

Linear or circular molecule, complexed with proteins, containing large numbers of genes.

Question 5

Requirements of RNA Polymerase?

Answer 5

Double stranded DNA template. Nucleotide building blocks in the form of NTPs (nucleoside triphosphates). High energy phosphate bonds in the triphosphates are broken to release energy for RNA synthesis.

Question 6

How does transcription occur?

Answer 6

RNA polymerase unwinds a short stretch of DNA to allow copying, following base pairing rules. RNA is synthesised in the 5’ to 3’ direction. Energy for DNA unwinding and RNA synthesis comes from the pyrophosphate release from nucleotide. RNA polymerase recognises promoter sequences (which show similarities in the -10 to -35 regions) and binds to these.

Question 7

What is pyrophosphate?

Answer 7

Oxygen and Phosphate

Question 8

How many nucleotides are synthesised per second by RNA Polymerase?

Answer 8

100.

Question 9

Can genes be simultaneously transcribed?

Answer 9

Yes, by multiple RNA Polymerases?

Question 10

What is required for transcription initiation?

Answer 10

Sigma subunit is needed for initiation. Gets released after the start of transcription.

Question 11

What signals the end of translation?

Answer 11

A hairpin loop termination signal due to complementary base pairing.

Question 12

Why is transcription regulated?

Answer 12

It requires a lot of energy, so makes sense to conserve energy by not producing certain proteins in different environments or cells.

Question 13

What is an inducer?

Answer 13

They disable repressors. The gene is expressed because an inducer binds to the repressor. The binding of the inducer to the repressor prevents the repressor from binding to the operator. RNA polymerase can then begin to transcribe operon genes.

Question 14

What is a transcriptional activator?

Answer 14

Protein (transcription factor) that increases gene transcription of a gene. Most are DNA binding proteins that bind to enhancers or promoter-proximal elements.

Question 15

Example of an activator?

Answer 15

The catabolite activator protein (CAP) activates transcription at the lac operon of E.Coli. cAMP is produced during glucose stavation. Binds to CAP, causing a conformational change that allows CAP to bind to a DNA site adjacent to the lac promoter. CAP then makes a direct protein-protein interaction with RNA polymerase that recruits RNA polymerase to the lac promoter.

Question 16

Example of an inducer?

Answer 16

Allolactose & lac operon. If lactose is present in the medium then some of it will get converted to allolactose by beta-galactosidase. Allolactose binds to the repressor and decreases the repressor’s affinity for the operator site. When glucose is also available, the lac operon is repressed.

Question 17

What is a repressor?

Answer 17

A DNA/RNA-binding protein that inhibits the expression of genes by binding to the operator or associated silencers.

Question 18

How does a DNA binding repressor work?

Answer 18

Blocks the attachment of RNA polymerase to the promoter, preventing transcription.

Question 19

How does a RNA binding repressor work?

Answer 19

Binds to mRNA and prevents translation of the mRNA into a protein.

Question 20

What is an operator?

Answer 20

A segment of DNA to which a transcription factor binds to regulate gene expression. Primarily occur in prokaryotes.

Question 21

What is an enhancer?

Answer 21

Short region of DNA that can be bound by proteins (activators) to increase the likelihood that transcription of a particular gene will occur. Found in both prokaryotes and eukaryotes.

Question 22

Example of a repressor?

Answer 22

Lac operon. LaxZYA transcribes the proteins needed for lactose breakdown. Lacl synthesises the repressor of this gene. The gene Lacl is situated immediately upstream of lacZYA but is transcribed from a different promoter. The lacZYA repressor is constituitively expressed. Always bound to the operator region which interferes with the ability of RNA polymerase to begin transcription.

Question 23

What is a constitutive gene?

Answer 23

A gene that is transcribed at a relatively constant level regardless of the cell environmental conditions.

Question 24

What is a facultative gene?

Answer 24

Transcribed only when needed.

Question 25

Operon theory?

Answer 25

Several genes are controlled by an operon. Forms one mRNA strand.

Question 26

Features of genetic code?

Answer 26

Triplet, non-overlapping. 1-6 codons can meant the same amino acid. 3rd position can often be unimportant. Stop codons mean the end of translation. Can be suppressed by tRNA mutations. Almost universal. Allows genes from one organism to be expressed in another.

Question 27

Why is the genetic code degenerate?

Answer 27

There are many instances in which different codons specify the same amino acid.

Question 28

How many binding sites do ribosomes have?

Answer 28

3 for tRNAs

Question 29

What do ribosomes require for protein synthesis?

Answer 29

mRNA amino acyl tRNAs to bring amino acids to the ribosome GTP - energy for synthesis Additional proteins - initiation factors, elongation factors and termination factors.

Question 30

Structure of tRNA?

Answer 30

Shape of a 3 leaf clover and has an anticodon as well as an "amino acid arm" where the complementary amino acid binds.

Question 31

How does tRNA get charged up?

Answer 31

1. ATP and the corresponding amino acid bind to the synthetase, releasing two inorganic phosphate molecules. Amino Acid + ATP -\> Aminoacyl-AMP + PPi 2. Binds the appropriate tRNA molecule's D arm and the amino acid is transferred. Aminoacyl-AMP + tRNA -\> Aminoacyl-tRNA + AMP Catalysed by a specific aminoacyl-tRNA synthetase. Has recognition sites for both tRNA and the specific amino acid.

Question 32

Steps of protein synthesis?

Answer 32

1. **Initiation** Small ribosomal subunit combines with mRNA and initiator tRNA. Large subunit joins. Special initiator tRNA is ued for recognising the start codon (Met) whilst different met-tRNAs are used for recognising internal AUGs. 2. **Elongation** Internal codons are translated in a series of translocation steps. rRNA catalyses the generation of a peptidyl bond between amino acids. The direction of synthesis is 5' to 3'. 3. **Termination** Synthesis stops when stop/termination/nonsense codons are encountered. The large and small subunits dissociate. tRNAs can't recognise stop codons, instead they are recognised by protein release factors. Following release factor association, polypeptide is released and the ribosome is recycled for next translation.

Question 33

Define polysome/polyribosome?

Answer 33

Number of ribosomes translating the same mRNA molecule form a polysome.

Question 34

Energy requirements for translation?

Answer 34

1. tRNA charging requires 1 ATP per aminoacyl-tRNA 2. initiation requires 1 GTP 3. termination requires 1 GTP 4. elongation requires 2 GTP per amino acid incorporated

Question 35

Differences between eukaryotic and bacterial protein synthesis?

Answer 35

1. Eukaryotic ribosomes are larger. 2. Some antibiotics differentially attack bacterial and eukaryotic protein synthesis 3. In bacteria there can be simultaneous transcription and translation due to no nucleus 4. Initiator amino acid in eukaryotes is methionine, in bacteria it is N-formyl methionine

Question 36

Examples of antibiotics that are used to attack bacterial translation?

Answer 36

1. Edeine hinders mRNA progression 2. Tetracycline prevents a-site tRNA binding 3. Erythromycin blocks protein exit from ribosome 4. Clindamycin blocks peptide bond formation