The genetic code - L8

Question 1

Describe the structure of tRNA.

Answer 1

• tRNAs have a cloverleaf secondary structure (The 5’ and 3’ ends are drawn together).
• tRNAs are folded into an L-shape - Coaxial stacking of helices and base-pairing between the ends of the D and TYC loops produce a flat, L-shaped molecule.

Question 2

How was the genetic code determined?

Answer 2

In vitro translation…
E.coli cell extract:

• isolate cell extract
• degrade mRNA with RNase
• inactivate RNase.
• add RNA and amino acids (one is 14C-labelled)
• precipitate protein & collect by filter binding.

Then filter to separate the proteins with which has radioactivity from luisine and radioactivity haven’t.

The first experiments by Marshall Nirenberg showed that poly(U) RNA specifically directed the incorporation of phenylalanine into protein, showing that the triplet UUU encodes phenylalanine.

Question 3

tRNAs can base-pair with more than one codon due to “wobble”:
What does ‘wobble’ mean - Non-Watson-Cricks base pairing at the 3rd base?

Answer 3

Many tRNAs have a modified nucleotide at the 1st position of the anticodon (the sequence within tRNA that base-pairs with the codon with an mRNA). G in the 1st position can recognise codons ending with C or U.
Inosine base-pairs with A, C or U (but not G). tRNALeu(IAG) can recognise CUA, CUC, CUU and UUA codons.

Question 4

What is involved in the process of tRNA aminoacylation?

Answer 4

tRNA aminoacylation is a two step process, involving formation of an aminoacyl-adenylate intermediate. AMP is generated.