Becker's World of the Cell 9th Edition by Lewis Kleinsmith, Jeff Hardin, Gregory Paul Bertoni

Exercise 11

Copolymer Analysis. In their initial attempts to determine codon assignments, Nirenberg and Matthei first used RNA homopolymers and then used RNA copolymers synthesized by the enzyme polynucleotide phosphorylase. This enzyme adds nucleotides randomly to the growing chain but in proportion to their presence in the incubation mixture. By varying the ratio of precursor molecules in the synthesis of copolymers, Nirenberg and Matthei were able to deduce base compositions (but usually not actual sequences)of the codons that code for various amino acids. Suppose you carry out two polynucleotide phosphorylase incubations, with UTP and CTP present in both but in different ratios. In incubation A, the precursors are present in equal concentrations. In incubation B, there is three times as much UTP as CTP. The copolymers generated in both incubation mixtures are then used in a cell-free protein-synthesizing system, and the resulting polypeptides are analyzed for amino acid composition.
(a)What are the eight possible codons represented by the nucleotide sequences of the resulting copolymers in both incubation mixtures What amino acids do these codons code for
(b)For every 64 codons in the copolymer formed in incubation A, how many of each of the 8 possible codons would you expect on average How many for incubation B
(c)What can you say about the expected frequency of occurrence of the possible amino acids in the polypeptides obtained upon translation of the copolymers from incubation A What about the polypeptides that result from translation of the incubation B copolymers
(d)Explain what sort of information can be obtained by this technique.
(e)Would it be possible by this technique to determine that codons with 2 Us and 1 C code for phenylalanine, leucine, and serine Why or why not
(f)Would it be possible by this technique to decide which of the three codons with 2 Us and 1 C (UUC, UCU, CUU)correspond to each of the three amino acids mentioned in part e Why or why not
(g)Suggest a way to assign the three codons of part f to the appropriate amino acids of part e.