The active site residues in some proteins seem to be "ultimately conserved," meaning that any change to that residue results in a complete lack of activity. Is it possible for the gene coding for that enzyme to evolve? Explain your answer.
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- 6. Riboswitches are RNA sequences found in the 5' untranslated region of messenger RNA (mRNA) that control gene expression in response to small molecule binding. These regulatory elements adopt two conformations: an "on" state when a specific small molecule ligand is bound and an "off" state in the absence of ligand. When the mRNA is "off", the gene remains untranslated. The structure of a guanine-responsive riboswitch was solved in 2004 (Nature, 432, 411-415) and is shown below at low (left) and high (right) resolution with a guanine derivative bound: (see Blackboard for color image) Guanine riboswitch with a guanine derivative (red) bound; zoomed out. U22 U47 HX U51 C74 Guanine riboswitch with a guanine derivative (HX) bound; atomic detail. Note: blue = nitrogen, red = oxygen. C and U follow standard nucleotide naming conventions. a. What interactions are occurring between the riboswitch and the guanine derivative that make binding favorable? b. Of these interactions, which ones…6. Similar to the class notes (Intro to Genetics), a segment of DNA (shown below) contains a promoter segment (the first 9 base pairs), a ribosome binding segment (the next 6 base pairs), and a segment that codes for protein synthesis which is started by the rest of the base pairs. ACTCCATTGAACCATTTCTATGATCCGCTAACG-... TGAGGTAACTTGGTAAAGATACTAGGCGATTGC-... A. When the DNA is induced to be copied to mRNA, the top strand is coding, meaning that the mRNA makes an identical copy of the lower strand (replacing T with U) The mRNA copy starts with the ribosome binding sequence. What is the sequence of the mRNA that will go to the ribosomes? B. What are the first 6 amino acids of the protein that are coded for by the mRNA? C. What would the amino acid sequence be if... i. a transition mutation occurred on the final G in the mRNA? ii. all of the G & C bases in the protein synthesis portion had transition mutations? iii. a point deletion mutation occurred in the ATA sequence (in the lower strand…1. You are studying an E. coli gene that specifies a protein. A part of its sequence is -Ala-Pro-Trp-Ser-Glu-Lys-Cys-His You recover two mutants for this gene that show no enzymatic activity. By isolating the mutant enzyme products, you find the following sequences: Mutant 1: Ala-Pro-Trp-Arg-Glu-Lys-Cys-His Mutant 2: Ala-Pro- What is the molecular basis for mutation 1 and for mutation 2? What is the DNA sequence that specifies this part of the protein? Wild-type amino acid sequence: NH3-Ala-Pro-Trp-Ser-Glu-Lys-Cys-His-COOH Mutant 1: NH3-Ala-Pro-Trp-Arg-Glu-Lys-Cys-His-COOH Mutant 2: NH3-Ala-Pro-
- . While perusing the E. coli K12 genome sequence,you come across a gene with no known function. Theamino acid sequence of the gene’s protein productshows weak similarities with known porins, proteinsthat cross a cellular membrane to let molecules suchas amino acid or sugar nutrients (or drugs like penicillin) pass through. Some porins are nonspecific and letany solute up to a certain size transit into the cell.Other porins are specific and allow the transit ofcertain sugars but not others. What genetic experiments could you do to try to determine whether thisnew gene has a specific function in allowing bacterialcells to scavenge the sugar maltose from the environment? Describe scenarios that might complicate yourexperimental approach.6. In the following graph, the dashed line shows the level of mRNA for a certain protein, Prot6, at various positions along the anterior-posterior axis of an insect embryo. The solid lines portray the levels of two regulatory proteins, A and B, that control translation of the Prot6 mRNA. A В Position Biology: How Life Works, Second Edition © 2016 W.H. Freeman and Company If both A and B stimulate translation of the Prot6 mRNA, then which graph, from the ones shown below, best approximates the expected level of Prot6 across the embryo? Briefly justify your answer. M K Q Position Position Position Position Position Protein level MRNA level2. Cyanobacteria are photosynthetic prokaryotes that are likely responsible for generating the oxygen- rich atmosphere we enjoy on Earth. Many species of cyanobacteria display "circadian rhythms" that manifest as oscillations in gene expression with a period of ~24 hours. The circadian rhythms are governed by a biochemical cascade that regulates the phosphorylation of a single protein. Remarkably, this circadian "clock" can be reconstituted in vitro with only 3 proteins, known as KaiA, KaiB and Kaic. Upon addition of Mg2+ and ATP to the reaction, KaiC cycles between being phosphorylated and dephosphorylated and reproduces the 24 hour period in the absence of any light cues. In vivo in the cyanobacteria, the phosphorylation of KaiC peaks at night, and this results in the activation of genes encoding components of the oxidative pentose phosphate pathway. Daytime photosynthesis m PSI ROS KaiA KaiB KaiC NADPH CO₂ fixation and glycogen synthesis 60 +ATP +Mg w 48 72 %P-KaiC 24 Detoxification…
- 1. A goal of synthetic biology is to genetically encode enzymes that have novel properties and enable them to carry out new chemical reactions. One way to encode new functional properties is to extend the genetic code beyond the naturally encoded 20 amino acids. To this end, a biotech company has engineered a bacterial species to encode an additional amino acid called "synthine", with a side chain with properties that are still a trade secret. In order for synthine to be incorporated into central metabolism, researchers at the company also used directed evolution to generate an enzyme known as synthine aminotransferase. (a) What reaction does synthine aminotransferase catalyze? What are the precursors and products? Why is it described as a "ping-pong" reaction? (b) The active site of synthine aminotransferase is shown in the protein structure image. What is the name of the prosthetic group? In the image on the left, the prosthetic group is covalently linked to the en- zyme. What is the…1. Given is the 30 nucleotides in the human gene for hemoglobin (the oxygen-carrying protein in the red blood cells): 3’ TAC-CAC-GTG-GAC-TGA-GGA-CTC-CTC-TTC-AGA 5’ a. What is the amino acid sequence based on this mRNA? b. A very important mutation in human hemoglobin occurs in this DNA sequence, where the T at nucleotide 20 is replace with an A. The mutant hemoglobin is called sickle cell hemoglobin and is associated with severe anemia. What is the amino acid replacement that results in sickle-cell hemoglobin?6. Different breeds of Korean cattle produce meat with varying amounts of stearic, linoleic and oleic acid. These differences are heritable and can be used in breeding programs to produce better beef. Mutations in fatty acid synthase are responsible for at least some of these differences. Which domain(s) would you expect to carry mutations in order to affect the ratios of stearic, linoleic and oleic acid produced? Justify your choice(s).
- 2. Cyanobacteria are photosynthetic prokaryotes that are likely responsible for generating the oxygen-rich atmosphere we enjoy on Earth. Many species of cyanobacteria display "circadian rhythms" that manifest as oscillations in gene expression with a period of -24 hours. The circadian rhythms are governed by a biochemical cascade that regulates the phosphorylation of a single protein. Remarkably, this circadian "clock" can be reconstituted in vitro with only 3 proteins, known as KaiA, KaiB and KaiC. Upon addition of Mg2+ and ATP to the reaction, Kaic cycles between being phosphorylated and dephosphorylated and reproduces the 24 hour period in the absence of any light cues. In vivo in the cyanobacteria, the phosphorylation of KaiC peaks at night, and this results in the activation of genes encoding components of the oxidative pentose phosphate pathway. Daytime photosynthesis PSI ROS KaiA KaiB Kaic NADPH 80 ROS co, fixation and Blycogen synthesis +ATP 60 Detoxification +Mg mechanisms 40…5. A mutant strain of Salmonella bacteria carries a mutation of the rho protein t hat has full activity at 37°C but is completely inactivated when the mutant strain is grown at 40°C. (Question # 21; Chapter 8-Genetics: An Integrated Approach). Speculate about the kind of differences you would expect to see if you compared a broad spectrum of mRNAs from the mutant strain grown at 37°C and the same spectrum of mRNAs from the strain when grown at 40°C. Are all mRNAs affected by the rho protein mutation in the same way? Why or why not?10. In eukaryotic cells, glycosylation activity of membrane proteins is found in the lumen of the endoplasmic reticulum (ER) and is accomplished by the enzyme oligosaccharyl transferase (OST), which adds oligosaccharides to the amino group of Asn residues of the consensus sequence Asn-X-Thr/Ser. a. The following ER membrane protein construct has a cytosolic N-terminal region and a membrane-spanning variable sequence. The variable sequence is a signal anchor (SA) sequence. Would you expect the C-terminal region of the B subunit to be glycosylated? Why or why not? variable sequence C-terminal region of B-subunit N-terminal region b. An additional transmembrane region is added to the construct prior to the variable sequence, as shown below. The transmembrane is a signal anchor (SA) sequence, while the variable sequence is a stop transfer anchor (STA) sequence. Would you expect the C-terminal region of the B subunit to be glycosylated? Why or why not? C-terminal region of B-subunit…