202 Biology Fall 2007 Homework 4 Due: September 20 or 21
NAME: T.A.: Discussion Section #:
1. (2 points) Many effective antibiotics affect bacteria by inhibiting translation. The experimental use of antibiotics has also been useful in determining the steps in protein synthesis. If you have an artificial mRNA with the sequence: AUGUUUUUUUUUUUUUAG, It will produce the protein: NH2-Met-Phe-Phe-Phe-Phe-COOH In your search for new antibiotics, you find one called putyermycin, which blocks protein synthesis. When you try to translate your artificial mRNA, the product is Met-Phe. What step in protein synthesis does putyermycin affect? Explain your answer. The antibiotic inhibits elongation. The polypeptide is made up of two amino acids and so they antibiotic is interfering with the translocation of the ribosome to the next codon of mRNA. 2. (1 point) a) What is the function of the Shine-Dalgarno sequence in prokaryotic mRNAs? The function of the Shine-Dalgarno sequence is to orient the ribosome to the proper AUG start codon. The rRNA molecule of the small subunit base pairs with this sequence and stabilizes the ribosome. Shorter answer is fine "site on the mRNA that the ribosome binds to for initiation of translation". b) What effect does the deletion of the Shine-Dalgarno sequence from an mRNA have on transcription and translation? There will be no effect on transcription. The Shine-Dalgarno sequence is not required for initiation of transcription by RNA polymerase. Deletion of the Shine Dalgarno sequence will affect translation. The ribosome requires the sequence for recognition of the correct AUG codon. The ribosome will not be stabilized to the mRNA without this sequence. 3. (5 points) The following is a diagram of pre-mRNA and its corresponding mature mRNA of a eukaryotic gene composed of 3 exons and 2 introns.
Pre-mRNA 2 3 4 5 6 7 8
mRNA
1
2
3
5
7
8
-AAAAAAAAAAAA
a) Which segment (1 to 8) represents the 5'cap? 1
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b) Which segment represents the 5' UTR? 2 c) Which segment represents the 3' UTR? 8 d) Which segments the represent exons? 3, 5, 7 e) Which segments represent the introns? 4, 6 f) In which segment would a single nucleotide insertion be most detrimental to the function of the resulting protein? 3 g) In which segment would a nonsense mutation be most detrimental? 3 h) In which segment does the start codon occur? 3 i) At the end of which segment does the stop codon occur? 7 j) Where would you expect to find the TATA box? The TATA box will not be in any of these segments. It is not part of the transcribed mRNA. 4. (1 point) A mutant bacterial cell has a defective aminoacyl synthetase that attaches a lysine to tRNA with anticodon AAA instead of a phenylalanine. What are the consequences of this mutation for the cell? At any point where an AAA codon specifies phenylalanine, lysine will be incorporated into the polypeptide. It is important to note that not all phenylalanine residues will be replaced by lysine. Only those encoded by the AAA codon will be replaced. This answer should be considered incorrect and only given partial credit. 5 (1 point) A nonsense mutation in the pathway for making the amino acid histidine prevents a strain of E. coli from growing on minimal medium. A mutation in the anticodon of a certain tRNA gene makes the anticodon complement the stop codon created by the original nonsense mutation. This tRNA mutation allows the E. coli to grow on minimal medium, however, it does not grow as well as wild type E. coli. Suggest a possible reason for why the tRNA mutation did not restore full growth. A mutation in a tRNA anticodon allows the tRNA to base pair with the stop codon and an amino acid will be incorporated into the polypeptide (rather than the synthesis terminating). This produces a functional enzyme which allows for growth. However, full growth is not restored because the incorporated amino acid is not the one originally coded for in the gene. Alternative answer: Some authentic stop codons would fail to terminate translation and this would result in proteins that are longer than the proteins should be.
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