Transcription And Translation

Question 1

Prokaryotic gene structure

Answer 1

Polycistronic = Promoter regulates transcription of several genes at a time
*can do transcription and translation at same time (all done in same location)

Question 2

Eukaryotic gene structure

Answer 2

Monocistronic = can only transcribe one gene at a time

• each gene has own promoter
• enhancer- for binding transcription factors

Question 3

Transcription

Answer 3

1. Initiation
2. Elongation
3. Termination

Question 4

Transcription Initiation

Answer 4

1. Prokaryotic
   * sigma factor finds -55 promoter region
   * RNA polymerase II haloenzyme -> closed complex
   * open complex -> short RNA made (thrown out)
   * sigma factor released -> RNA polymerase II continues
2. Eukaryotic
   * TFIID binds TATA box -> TFIIB binds -> connects RNA polymerase II
   * preinitiation complex forms (closed)
   * complex opens -> elongation

Question 5

Transcription Elongation

Answer 5

1. Prokaryotic
   * RNA polymerase subunits (sigma= initiator, core enzyme 2 alpha and 2 beta)
   * RNA made 5’->3’
   * template strand (antisense) 3’->5’
   * mRNA= coding (sense) strand w/ U instead of T
2. Eukaryotic
   * RNA polymerase (I = rRNA, II = mRNA, III = tRNA)
   * process same as prokaryotes

Question 6

Transcription termination

Answer 6

1. Prokaryotic
   A. Rho-independent: spontaneous, new RNA forms hairpin
   B. Rho-dependent: requires protein Rho - separate DNA and RNA
2. Eukaryotic
   * PolyA polymerase signal
   • poly-A tail transcribed
   • termination factors: free RNA a few kB from poly-A tail
     
     • CPSF, CstF

Question 7

Transcription inhibition

Answer 7

1. Prokaryotic
   * Rifampin- antibiotic for tuberculosis
   
   • binds to bacterial RNA polymerase -> chain stops after 3 nts
     * Dactinomycin- intercalated between DNA bases and inhibits initiation/elongation
   • not prokaryote/eukaryote specific
   • antibiotic and anti tumor
2. Eukaryotic
   * alpha-amamitin: poison in mushrooms
   
   • inhibits RNA polymerase II
   • symptoms: GI disturbances, electrolyte imbalance, fever, liver and kidney dysfunction
   • death w/in 10 days 10-20% of pts.
   • no antidote

Question 8

Streptomycin

Answer 8

Inhibits translation initiation by distorting the 30S subunit of ribosomes

Question 9

Tetracycline

Answer 9

Inhibit translation elongation by blocking the A site which doesn’t allow aminoacyl-tRNA in

Question 10

Chloramphenicol

Answer 10

Inhibits prokaryotic peptidyltransferase

*high levels can inhibit mitochondrial protein synthesis

Question 11

Puromycin

Answer 11

Structurally similar to aminoacyl-tRNA and accepts peptide from P site -> premature termination
*prokaryotes and eukaryotes

Question 12

Erythromycin

Answer 12

Inhibits translocation by binding to the 50S subunit and blocking the tunnel for the polypeptide to leave the ribosome

Question 13

Translation initiation

Answer 13

1. Initiation factors (IF) form complex (30S) -> tRNA bought to A site
2. GTP on IF-2 hydrolyzed -> 50S joins 30S -> 70S complex -> IFs released

Question 14

Translation elongation

Answer 14

1. Elongation factor (EF) brings tRNA to codon in A site -> GTP -> GDP
2. Peptidyltransferase catalyze peptide bond formation -> chain moves P -> A site
3. MRNA shifts in ribosome sites (5’->3’) = translocation
   * EF-G-GTP, GTP -> GDP

Question 15

Translation Termination

Answer 15

Termination codon recognized -> polypeptide released

1. Prokaryotic
   * Shine-Delgarno sequence
   * 3 IFs
   * 1st aa = formulated methionine (fMet)
2. Eukaryotic
   * 5’ cap directs binding
   * many IFs

Question 16

Protein folding

Answer 16

1. Spontaneous
   * many proteins
   * require suitable physiological conditions
2. Chaperone-assisted
   * Heat-shock proteins (Hsp) and chaperonins
   * fxn as molecular chaperones
   * require ATP

Question 17

Proteosomal degradation

Answer 17

• selectively degrade damaged/short-lived proteins
• ubiquitin modification to target
• requires ATP

Question 18

Misfolded proteins

Answer 18

Cause amyloid fibrils

• Brain: Alzheimer’s, Parkinson’s, mad cow disease, Huntington’s
• Pancreas: type II diabetes

Question 19

Protein processing and targeting

Answer 19

1. Carb addition
2. Lipid addition
3. Mannose-6-P: signal for proteins to go to lysosomes

Question 20

I-cell disease

Answer 20

Caused by protein mistargeting

• dec. in enzyme that phosphorylates C6 on mannose
• Characteristics:
  
  1. Skeletal abnormalities
  2. Restricted joint movement
  3. Coarse facial features
  4. Severe psychomotor impairment
• death in early childhood
• very rare
• autosomal recessive