You have isolated an inhibitor, named edeine, from an undrinkable batch of home-

Question

You have isolated an inhibitor, named edeine, from an undrinkable batch of home-brewed beer that was contaminated by an unknown microbe. Edeine inhibits eukaryotic protein synthesis but has no effect on DNA or RNA synthesis. When added to rabbit reticulocyte protein-synthesis extract (see PDF), edeine stops hemoglobin synthesis after a short lag. In contrast, the translation-elongation inhibitor cycloheximide stops protein synthesis immediately. Analysis of the edeine treated system showed no polyribosomes containing hemoglobin mRNA after protein synthesis stopped. Instead, all the globin mRNA accumulated in an unusual 46S peak. As expected, cycloheximide treatment results in stable polyribosomes containing hemoglobin.     

A. What step in protein synthesis does edeine inhibit?

B. Why is there a lag between addition of edeine and cessation of protein synthesis?

C. Explain whether polyribosomes would disappear if you added cycloheximide at the same time as edeine?

Explanation / Answer

A) During initiation of protein synthesis, ribosomal 40S subunits (Smaller subunit with eIF3 and the eIF2-GTP/Met-tRNAi complex, come near the 5? end of the capped mRNAs. The mRNA is scanned by the 40S subunit for the start codon, which is then positioned near the P site of 40S subunit. The eIF2 dissociates from the 40S subunit due to GTP hydrolysis. The 60S subunit will now associate with the 40S subunit in presence of eIF5B.

Edeine inhibits binding of aminoacyl-tRNAs to the P site of both 30S subunits and 70S ribosomes, as well as to eukaryotic 40S subunits. The inhibition of binding of aminoacyl tRNAs to P site is due to codon-anticodon misreading. Hence, fmet tRNA cannot bind. Further, the large subunit cannot bind to the smaller subunit. Thus, initiation of proteins synthesis is inhibited.

B) The gap between edeine effect and lack of protein synthesis is because of the ribosomes that have already initiated protein synthesis. These ribosomes will continue translating the protein as elongation step is not affected. The inhibitor is added after 2 min after translation has been initiated. Only new protein translation is inhibited by edeine. Hence, a gap will be observed before translation completely inhibits.

c) Cyclohexamide is an inhibitor that affects protein elongation during translation. It blocks the movement of peptidyl-tRNA from A site to the P site on ribosomes. This transfer requires Transferase II, TF-II, and GTP hydrolysis. Cyclohexamide traps polyribosomes on the mRNA. When both edeine and cyclohexamide are added simultaneously, the have already started elongation will be trapped in mRNA. Hence, these stalled polyribosomes will be seen that are trapped. Not all polyribosomes will therefore disappear. Addition of both edeine and cyclohexamide will inhibit both initiation and elongation. Hence, protein synthesis is completely abrogated.

Related Questions

Navigate

• Browse (All)
• Browse Y
• Subjects

Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.