MCDB131 Old Final

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University of California, Santa Barbara *

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131

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Biology

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Jan 9, 2024

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1. What is the proton gradient generated in the cell membrane used for? It powers nutrient uptake (LacY permease), compound export (AcrB), flagella (MotA/B), and generation of ATP (ATP Synthase) among other processes. 2. Transcription termination in E. coli requires the mRNA-binding protein Rho and rho utilization (rut) sites in 3' end of the transcript. False Transcription termination by RNA polymerase can occur through two mechanisms: (1) An attenuator stem loop structure formed by G-C base pairs, followed by a run of U's and (2) physical interaction with Rho, a protein that binds a growing mRNA transcript and disrupts the DNA-RNA helix at the RNAP active site. Thus, termination does not strictly require Rho. 3. The base excision repair pathway repairs what kind of lesions? Base excision repair is responsible for repairing small, non-helix distorting mutations from DNA such as: deaminated, oxidized, or alkylated bases (or the incorrect incorporation of uracil). 4. Which of these are false (there may be more than one)? A. The cell maintains a slightly underwound chromosome to facilitate strand opening for transcription. B. Sigma factor associates with RNA polymerase to direct it to specific operator sequences at the beginning of genes. C. The origin of replication is TA-rich to facilitate melting of the DNA. D. The dif site on the E. coli chromosome is recognized by the XerC integrase to resolve concatenated chromosomes. E. Type I topoisomerases use ATP hydrolysis to remove supercoils in DNA molecules. Sigma factor does associate with RNA polymerase to bind DNA, but it does not recognize operator sequences. Sigma binds the promoter elements on the DNA (-35, -10). Transcription factors, such as repressors, bind to operator sequences near the promoter to affect RNAP. Type I topoisomerases do not use ATP to remove supercoils; they will relax the DNA by making a nick in one strand when too many supercoils build up. Type II topoisomerases, however, can use ATP to introduce negative supercoils into the DNA by cutting both strands and passing the other side of the helix through the break (think: circle O --> figure 8). 5. Which DNA repair pathway(s) can remove pyrimidine dimers from the genome? Select All that apply: A. Nucleotide excision repair B. homologous recombination C. Base excision repair D. Photoreactivation. E. None of the above

6. During binary fission, each daughter cell will inherit one methylated and one unmethylated strand of DNA immediately following DNA replication and division. True 7. What happens to ribosomes synthesizing the leader peptide of the tryptophan operon when there are low levels of tryptophan in the cell and why? Which stress response uses a similar mechanism to detect starvation and how? When [Trp] is low, the ribosome stalls in the leader region of the Trp operon where there are successive Trp codons. Stalling leads to the formation of an anti-attenuation stem loop (loop 2- 3) since the ribosome is blocking region 1 from forming a loop with region 2. This means that RNA polymerase can continue making the Trp operon transcript, and gene expression occurs. The stringent response is a stress response that also relies on ribosome stalling to detect global amino acid depletion, rather than just tryptophan. Stalled ribosomes recruit RelA, a protein that synthesizes (p)ppGpp, which is an "alarmone" and can directly alter gene expression by interacting with RNA polymerase. The outcome of the stringent response is to slow or stop growth by pausing the synthesis of rRNA and tRNA. 8. (True/False) MreB is found in all rod-shaped bacteria. True 9. What do you predict the phenotype of a strain lacking the chromosomal dif site to be, when compared to the wild type parental strain? A small percentage of the population will be filamentous, since a small percentage of the time dividing cells might experience chromosomal concatenation. As a result of lacking the dif sequence (integration site for the XerCD integrases) to separate them, SlmA will prevent FtsZ from forming the division septum and the cell becomes filamentous. 10. Which of the following has the greatest affinity for its electrons? A. NADH B. Glucose C. H2O D. FADH2 Oxygen is the terminal electron acceptor of aerobic respiration, which is reduced to form water. This process will have the highest reduction potential, and thus the greatest affinity for electrons.

11. Which term best describes this reaction: NH4+ + 3O2 → 2NO2− + 2H2O + 4H+ ? A. Organoheterotrophy B. Fermentation C. Photoheterotrophy D. Lithotrophy E. Anaerobic respiration A lithotroph uses inorganic substrate to obtain reducing equivalents. Ammonia, in this example, serves this purpose, and is oxidized into nitrite. 12. What is the source of electrons for oxygenic photosynthesis? A. Oxygen B. Light C. Water D. Carbon dioxide 13. In which order will the following electron carriers act in an electron transport chain? Electron Carrier Reduction Potential (V) A -0.3 B 1.0 C -0.6 D 0.5 C -> A -> D -> B 14. A biofilm of Vibrio fischeri is mixed with Variovorax paradoxu , a bacterium that uses homoserine lactone as an electron donor in its metabolism. What effect will this have on the V. fischeri bacteria? The metabolizing bacterium will decrease the effective concentration of homoserine lactone in the biofilm, reducing levels of activation of the lux gene pathway, thus reducing fluorescence. Because homoserine lactone is used by V. fisheri as a quorum sensor, they will also exhibit gene expression patterns consistent will lower population densities. 15. Why is iron limiting in the environment? Describe the iron uptake pathways of Gram negative and Gram positive bacteria. Iron is either sequestered in iron oxides or it is insoluble. Gram negative bacteria use FepA in the outer membrane as a passive transporter, which can be actively powered using TonB/ExbBD. The siderophore is then bound by FepB in the periplasm and ferried to the FepDC ABC transporter.

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In Gram positive bacteria, a gradient of increasingly high affinity siderophores is constructed in the cell wall to generate net flow of iron towards the cell. These final siderophore complexes can be imported cells and degraded to release coordinated iron. 16. Colicin systems provide certain bacteria competitive advantages for nutrient acquisition. A zone of clearance assay is common for testing whether a colicin is secreted into the media, as colicin activity will kill neighboring bacterial cells that lack a cognate immunity protein. There are three genes involved in colicin E synthesis, cea (colicin E) , cei (Immunity) , and cel (lysis protein) and they are regulated by PSOS. Predict the outcomes of the following Colicin E expression in these genetic backgrounds along with an SOS response inducer (mitomycin C). Genetic background Mitomycin C Expression of Colicin E Presence of Zone of clearance Wild-type - No No Wild-type + Yes Yes Δ cel + Yes No Δ recA + No No Δ cel - No No Δ recA - No No 17. Two genes are identical in their sequence and regulation except for their Shine-Dalgarno sequence. Gene 1 has perfect complementation to the Anti Shine-Dalgarno, Gene 2 has two substitutions of T -> C. Which of these genes do you predict will have more ribosomes on the polysome? Why? Gene 1 would have more ribosomes on its transcript because its Shine Delgarno has much higher affinity for the 16S of the ribosome (the anti-Shine Delgarno). 18. How can a small RNA be used to reduce the expression of a gene? How could its expression be regulated intrinsically? sRNA's can compete for ribosome binding at the Shine Delgarno sequence, thereby preventing translation. They can also decrease stability of a gene transcript, causing it to be degraded more quickly by exonucleases. The mRNA itself could form secondary structures at prevent ribosome binding at the ribosome binding site without the help of a sRNA. 19. Which of the following statements best describes Wolbachia infection? A. Infected male wasps can only mate with non-infected female wasps B. Infected male wasps prefer females infected with the same Wolbachia strain C. Two wasps infected with different strains of Wolbachia can mate

D. Wolbachia is a female-killing bacteria. 20. Which of the following statements is true regarding Type V and Type VI secretion systems? A. Type VI requires an exporter protein encoded by a gene called cdiB B. Type VI requires a receptor protein on the target cells C. Type V requires a receptor protein on the target cells D. Type V tip is composed of VgrG/PAAR 21. Which of the following correctly describes a microscopy technique? A. Scanning electron microscopy illustrates internal structures of a sample B. Fluorescence microscopy visualizes structures by using pigments and measuring their absorption of visible light C. Electron cryotomography provides high resolution imaging of dynamic biological processes in real time D. Transmission electron microscopy generates 3D images of surfaces by passing electrons through a sample E. None of the above 22. A bacterial culture is enumerated using two methods. First, the optical density is measured and compared to a standard curve for CFU/mL. Second, the culture is serially diluted followed by spreading on plates. Which method will likely give a higher CFU/mL measurement? Why? Optical density measures the absorbance of visible light by the bacterial culture and will be influenced by both living and dead cells. Serial dilution and plating will only measure viable cells (i.e. ones that can form colonies). Optical density will give a higher reading. 23. Gene X codes for an inducer which has been engineered to bind DNA in the presence of ligand Y. The recognition sequence for protein X is found in-between the -10 and +1 transcription sites for a sigma factor (sigma 51) which regulates a copy of GFP. Sigma 51 is also upregulated by cAMP. An additional factor, Z, induces a very strong repressor for gene X. Rank the GFP fluorescence from cells grown in the following cultures: A. LB B. LB + Y C. LB + Z D. LB + Y + lactose E. LB + glucose From most GFP to least GFP fluorescence: LB + Y + lactose > LB + Y > LB > LB + glucose >= LB + Z (Both glucose and factor Z would repress GFP expression, with the product of Z induction likely more potent repression since it is a locus specific repressor, but both conditions could be similar)

24. When a bacterium is returned to physiological temperature, what molecular methods are used to reduce the expression of heat shock genes? Sigma H mRNA folds into a secondary structure that inhibits ribosome binding and thus translation. Leaky expression of Sigma H may occur, but DnaJ/DnaK chaperones shunt any translated RpoH/Sigma H protein to degradation. 25. An Ames test is performed using an MNNG-soaked paper. What do you expect to see? A. The mutagen will prevent colonies from forming on the plate B. Colonies will form far away from the mutagen and will become increasingly smaller closer to the center C. Colonies will form close to the mutagen and will become increasingly smaller further away D. Colonies will be evenly spread out on the plate and of uniform size E. None of the above 26. Describe two methods by which a bacteria may defend itself from infection by a DNA virus. CRISPR-Cas Restriction-Modification system CRISPR-Cas encodes for "designer nucleases" which utilize ~20-30bp RNA to target complimentary and cleave foreign DNA. The restriction modification system is a set of enzymes which recognize and cut short (~6-7bp) palindromic sequences of DNA. The modification system typically use methylation to denote "self", as the restriction enzymes are then unable to bind that sequence. 27. Which of the following is true regarding horizontal gene transfer? A. Bacteria can only acquire new genes from transduction B. Bacteria can uptake RNA from the environment through a process called transfection C. Bacteria can uptake DNA from the environment through a process called transformation D. Bacteria can transfer genes to another bacteria with a pilus through a process called transduction 28. Which of the following is false regarding transcription and translation? A. The 5’ end of an mRNA contains a triphosphate B. EF-G-GTP is involved in ratcheting of the ribosome during elongation C. Tetracyclines block the A-site of the ribosome D. There are multiple RNA Polymerases for transcribing different genes E. Proteins are able to occupy the A-site of the ribosome

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29. How long will it take a bacterial population of 100 cells to reach 1600 cells, given a doubling time of 30 minutes? 120 minutes 30. Which of the following is true regarding iron uptake in bacteria? A. The genetic regulation of siderophore transporter is tied to the biosynthesis of the siderophore molecule. B. Siderophore affinity for iron may be so extreme that the chemical is degraded to free the iron C. ATP drives the uptake of siderophores across the outer membrane in gram negative bacteria D. The Ton-system is involved in the uptake of nutrients other than iron A - there are noted instances of bacteria possessing transport systems (e.g. FepAB) without producing the specific siderophore themselves (an example of iron piracy) B- Enterochelin is utilized to remove iron from other chelators and is ultimately so tightly bound that ferrienterobactin esterase is required to cleave the molecule and retrieve the iron. C- proton flux drives the conformational change of the TonB protein, which is a protein tethered to the inner membrane ExbB and ExbD. It serves to propagate conformational change to the outer membrane transporter as part of the uptake mechanism. This is necessary due to the periplasmic space lacking high energy molecules which are normally used to drive conformational changes (ATP for example) D- Ton system is involved in Vitamin B12 uptake as well. 31. Which of the following is true regarding negative staining? A. It is a differential staining technique to identify Gram-negative bacteria B. It is a simple staining technique to exclude Gram-positive bacteria C. It is a differential staining technique to add contrast to the surrounding environment D. It is a simple staining technique to stain the bacterial cell white 32. Which of the following is true? (Select all that apply): A. MinC is a DNA binding protein B. MinD has an FtsZ-inhibiting domain C. SlmA has both DNA binding and FtsZ-inhibiting domains D. The min system prevents septation at the chromosome E. The slm system prevents septation at the poles of cells 33. Which of the following is true regarding DNA replication? (select all that apply): A. xerC and xerD are essential genes B. NAD is used to ligate DNA fragments together C. Parent DNA strands are rejoined after replication

D. DNA polymerase III synthesizes Okazaki fragments E. Binding sites for Tus proteins are encoded more than 180 degrees away from the origin in each direction 34. Which is false regarding plasmid partitioning systems? A. High copy plasmids do not use plasmid partitioning systems B. Antitoxin proteins encoded in plasmid TA systems are more stable than their corresponding toxins C. ParM is an actin homolog that pushes plasmids to opposite sides of the cell D. ParR is a DNA binding protein E. None of the above are false 35. Which of the following is false regarding nutrient import? A. Energy is expended in the import of every nutrient B. A glycolysis intermediate begins the phosphorylation cascade utilized by phosphotransfer systems C. The Enzyme II CBA complex of the glucose importer performs catabolite repression when phosphorylated D. ABC transporters can both import and export compounds E. ATP is absent from the periplasm

MCDB131 Old Final

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