Cell Biology and Signalling

Question 1

What is the structure of a prokaryote?

Answer 1

No nucleus - DNA lies freely in the cell

Single circular DNA molecule (+ plasmids)

10⁶ - 5 x 10⁶ bp

No internal compartmentalisation

Organo-, photo- or lithotrophic

No cytoskeleton

0.1 - 1 μm

Question 2

What is the structure of a eukaryote?

Answer 2

Nucleus contains DNA

Multiple linear DNA molecules (chromosomes)
1.5 x 10⁷ - 5 x 10⁹ bp

Have a cytoskeleton

Have cytoplasmic organelles

10 - 100 μm

Question 3

What is the function of the nucleus, the nucleolus, the cytoplasm and the cytosol?

Answer 3

Nucleus → contains the genome, site of DNA and RNA synthesis, surrounded by a nuclear envelope.

Nucleolus → rRNA genes, rRNA, ribosomal proteins, ribosome subunit assembly.

Cytoplasm → contains organelles and cytosol

Cytosol → place for many essential chemical reactions and protein synthesis

Question 4

What is the function of the ER, the Golgi body, mitochondria and chloroplasts?

Answer 4

ER → membrane-enclosed labyrinthine space. Rough ER contains ribosomes are protein synthesis. ER also produces lipids and functions as Ca²⁺ storage.

Golgi body → stacks of flattened membrane, receives lipids and proteins from the ER, modifies them and dispatches them to other parts or the exterior of the cell

Mitochondria → site of aerobic respiration

Chloroplasts (in plants and algae only) → site of photosynthesis

Question 5

What is the function of lysosomes, endosomes and peroxisomes?

Answer 5

Lysosomes → contain digestive enzymes to degrade defunct organelles, macromolecules, as well as particles taken up from the cell exterior by endocytosis. Release nutrients from the breakdown of food particles.

Endosomes → Endocytosed material must pass through endosomes (vesicles) before being delivered to lysosomes.

Peroxisomes → contain enzymes for various oxidative reactions including H₂O₂​ to inactive various toxic molecules.

Question 6

How is the folding of RNA different to that of DNA?

Answer 6

DNA forms a double helix between the two complementary strands, which is progressively coiled up to form chromosomes.

RNA exists only as a single strand. Hydrogen bonds form between complementary base pairs in the strand, making the RNA molecule fold up onto itself.

More hydrogen bonds can then form across the backbone, between folded regions of the chain. These can be unconventional base-pair interactions, eg. A-G.

Question 7

How is DNA transcribed by RNA polymerase?

Answer 7

RNA polymerase unwinds the DNA double helix. It is only unwound locally, around 11 base pairs at a time.

Ribonucleoside triphosphates enter the active site of RNA polymerase via the ribonucleoside triphosphate tunnel.

RNA polymerase catalyses the formation of phosphodiester bonds between the RTPs. The newly synthesized RNA transcript then exits RNA polymerase.

The RNA produced is complementary to the DNA template strand, but it has the same code as the coding strand.

Question 8

How does transcription start in prokaryotes?

Answer 8

The sigma factor of RNA polymerase scans the DNA sequence for the promoter sequence, which contains recognition sequences at -35 and -10 base pairs upstream of the +1 site.

RNA polymerase then binds to the DNA double helix and transcription begins. The sigma factor is released, RNA polymerase clamps down on DNA and transcription continues.

Transcription stops when RNA polymerase reaches the stop sequence within the DNA code. RNA polymerase and the completed RNA strand are released, and the sigma factors rebinds to RNA polymerase.

Question 9

Can the DNA strand be transcribed in either direction?

Answer 9

Yes. The relative positions of the -35 and -10 sequences determine which direction RNA polymerase moves along the DNA strand.

RNA polymerase goes from - 35 to -10.

Question 10

How does transcription start in eukaryotes?

Answer 10

The TATA binding protein binds to the TATA box upstream of the transcription start site. This then recruits TF2D, then TF2B.

TF2E, TF2F and RNA polymerase II are all then recruited. TF2H is then recruited, which has helicase activity and unwinds the DNA double helix.

TF2H then phosphorylates RNA polymerase II, releasing the polymerase from most of the general transcription factors.

Transcription then continues until the termination sequence is reached.

Question 11

How is eukaryotic pre-mRNA modified after transcription?

Answer 11

The pre-mRNA strand has a methylguanosine cap added at the 5’ end. It also has a poly-A tail ranging from 150 to 250 base pairs in length added to the 3’ end.

pre-mRNA is modified in order to stabilise it, aid nuclear export, as well as act as a checkpoint for protein synthesis machinery.

pre-mRNA modification actually defines mRNA, because it is the only type of RNA to be modified.

Question 12

What is RNA splicing?

Answer 12

Eukaryotic pre-mRNA contains introns and exons. Introns are non-coding sequences, and so they must be removed in order for the transcribed protein to function properly.

There are 3 specific sequences within the intron that act as recognition sequences for the spliceosome.

Spliceosomes perform the splicing, and these are made up of small nuclear RNAs and proteins called snRNPs. Spliceosomes can act only any pre-mRNA strand that contains these 3 sequences.

Question 13

What are the potential advantages of having introns and exons?

Answer 13

pre-RNAs can undergo alternative RNA splicing to produce different mRNAa and, therefore, different proteins from the same gene.

Many exons encode specific domains, and ‘exon-shuffling’ between genes may have contributed to the evolution of new genes and proteins.

Question 14

How do tRNAs bind to amino acids?

Answer 14

The binding of an amino acid to its corresponding tRNA is catalysed by aminoacyl-tRNA synthetases.

When the amino acid and tRNA are about the be bound together, they are said to be ‘charging’.

A high-energy ester bond forms between them using the energy released from splitting ATP into AMP + 2Pi. The tRNA is now said to be charged.

Question 15

What is the structure of a ribosome?

Answer 15

The large ribosomal subunit consists of around 49 proteins and 3 rRNAs. It covalently links amino acids together.

The small ribosomal subunit consists of around 33 proteins and 1 rRNA. It matches the mRNA codon to its corresponding tRNA.

There are 3 sites within the ribosome:

Aminoacyl (A) site

Peptide (P) site

Exit (E) site

Question 16

What are the steps in translation?

Answer 16

1 → the charged tRNA binds to mRNA by forming base pairs with the correct nucleotide triplet.

2 → the amino acid in the P site is uncoupled from its tRNA and a covalent peptide bond is formed with the amino acid in the A site.

3 → The large subunit shifts relative to the small subunit. The tRNA in the P site moves to the E site, and the tRNA in the E site moves to the P site.

4 → The small subunit moves 3 nucleotides along the mRNA. The empty tRNA is ejected, and the empty A site is ready to receive the next charged tRNA.

5 → This process is repeated until a stop codon is reached.

Question 17

How does translation start and stop?

Answer 17

Start:

The small ribosomal subunit with translation initiation factors and the initiator tRNA bound to it bind to the mRNA strand. This moves along the mRNA strand searching for the first AUG.

At the start codon, the translation initiation factors dissociate and the large ribosomal subunit binds.

Translation then continues as normal.

Stop:

When the ribosome reaches a stop codon, a release factor binds to the A site.

Peptidyl transferase adds H₂O to the released polypeptide chain instead of an amino acid, forming the -COOH group.

Both ribosomal subunits dissociate, and translation has terminated.