Question

1. Some amino acids are post-translationally removed from the C-terminal end of the beta-lactamase enzyme from B. imaginarium (i.e. - after it is translated and released from the ribosome, a protease chews off some amino acids).  The wild-type enzyme, which has had the amino acids removed from the C’-terminus, is 246 amino acids in length and the C-terminal amino acids are shown below aligned with the C-terminal amino acids of a frameshift mutant, which – due to a frameshift mutation - is not processed at its C-terminus. The wildtype and frameshift mutant have identical coding sequences except for the insertion or deletion of a single nucleotide in the DNA encoding the C-terminal region of the protein.  Use this information to answer the questions below.                   

codon#: 243 244 245 246

wt: gly leu asn ser-COOH

mutant: gly his gln gln thr trp met asn his glu  gly-COOH

1. What was the mutational event that gave rise to the frameshift mutant?  (identify which codon contains the inserted or deleted nucleotide and the position within the codon where the mutation occurred.
2. Deduce the number of amino acids in the wild-type enzyme prior to processing of the C-terminus.
3. Deduce, as far as possible, the sequence of the C-terminus of the wild-type enzyme prior to processing (you will not get a unique answer due to the degeneracy of the genetic code).

Answer 1

A) CUU, CUA, CUC, CUG codes for leucine, the 244 th amino acid in the normal protein is Leu, in the mutant protein it is changed to histidine, this is due to the insertion of a single nucleotide, the 244th codon was CUC, A is inserted after C, so the codon has changed to CUA, then codon begins with C, it can be either CAA or CAG.which codes for glutamine, the 245 th amino acid was Asn it is coded by AAU, AAC so the 245 the codon in the mutant mRNA is CAA which codes for histidine.

the mutation is due to the insertion of A after first C in the 244th codon.

B) total number of amino acids in the wildtype protein is 246. ( number of codons till the last amino acid= number of amino acids, each codon codes for one amino acid except the stop codon)

C)one possible mRNA that codes for the C terminus of the protein is  GGG-CUC-AAC-AGU

the DNA sequence is 5`-GGG-CTC-AAC-AGT-3`

( both coding sequence and mRNA has same sequence but in DNA U`s are replaced with Ts)