Yeast is a haploid organism. You are given a yeast his auxotroph, his-1 This aux
ID: 167484 • Letter: Y
Question
Yeast is a haploid organism. You are given a yeast his auxotroph, his-1 This auxotroph cannot make an amino acid histidine, which is essential for yeast's growth. So, this auxotroph does not grow on minimal media plates. Molecular analysis of this his-1 auxotroph shows that this mutation is caused by a nonsense mutation, which changes tyrosine codon, UAC, to a stop codon, UAA near the beginning of the his-1^+ gene. After a large-scale mutagenesis of this his-1 auxotroph, you have isolated a few pseudorevertants with nonsense suppressor genes. Some of them grew as good as prototrophs on minimal media plates. Some of them grew on minimal media plates, but not as good as prototrophs. Explain this results by listing all possible genes to be mutated to become nonsense suppressor mutations. You isolated a recessive (loss-of-function) mutation of a membrane-spanning enzyme in Drosophila. Since this mutation is viable, you examined the presence of the protein of this gene with western blotting. The mutant flies had the same amount as WT flies. Next, you measured the activity of this enzyme. Surprisingly the mutant showed the same activity as WT flies However, the mutant form of the enzyme was found in the cytoplasm, instead of in the plasma membrane. What is a possible molecular lesion of this mutation?
Explanation / Answer
Q.No 1
Anticodon mutations conferring nonsense suppression can beidentified in yeast sup70 gene and this gene codes for glutamine tRNA (CAG). Mostly amber suppressor glutamine tRNA(UAG) can be translated 5’-CAG-3’ glutamine codons . A-C mismatching takes place in the first position of the specific with low efficiency. Yeast glutamine tRNA suppressors express genes in the 5’-UAG-3’ and 5’-UAA-3’ codons. Transformation of nonsense mutant SUP45 gene encoding wild type tRNA. Overexpression of these genes leads to the increase the length eRF1 protein and it can compensate non-sense mutants sensitivity to relatively high temperature.
Q.No 2
Mutations in the active site of enzymes may result in the inactive enzyme substrate complexes when the catalytically inactive enzyme is overproduced. At the same time, inhibitory proteins are isolated by deletion mutations or point mutations. These mutations can express full length as such in the wild type proteins. This mutation changes location of the enzyme from plasma membrane to cytoplasm due to production of truncated proteins.
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