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In the following procedure, the protein of interest, CA, is…

In the following procedure, the protein of interest, CA, is initially concentrated by precipitation from a 30% saturated ammonium sulfate solution. If you were interested in using ammonium sulfate precipitation to isolate a highly charged peptide, would you expect the peptide to precipitate from a low or high concentration ammonium sulfate in solution? Briefly justify your answer.   Procedure from the reference cited below: “Briefly, cells expressing the wild-type CA protein were lysed through a microfluidizer. Soluble CA protein in the clarified lysate was concentrated by precipitation from 30% saturated ammonium sulfate. CA protein was redissolved in 50 mM Tris (pH 8.0) and functionally purified by the addition of sodium chloride to a final concentration of 2.5 M. After two rounds of functional purification, the wild-type CA protein was resuspended in 50 mM sodium phosphate buffer (pH 7.5) and dialyzed against the same buffer. The dialyzed sample was further purified by a subtractive anion exchange chromatography step using a Q-HP HiTrap column (catalog no. 17-1154-01, GE Healthcare, Piscataway, NJ). Purification of 2Mut and 4Mut CA mutants followed the same protocol that was used for wild-type CA except that 200 mM β-mercaptoethanol was included in all buffers throughout the process. The functionally purified capsid mutant proteins were redissolved in 50 mM Tris (pH 7.5) and 40 mM β-mercaptoethanol prior to dialysis in the same buffer, and a subsequent purification with subtractive anion exchange chromatography was performed.” Tsiang, M., Niedziela-Majka, A., Hung, M., Jin, D., Hu, E., Yant, S., Samuel, D., Liu, X., & Sakowicz, R. (2012). A Trimer of Dimers Is the Basic Building Block for Human Immunodeficiency Virus-1 Capsid Assembly. Biochemistry, 51(22), 4416-4428.

This figure shows the results of a two-dimensional gel that…

This figure shows the results of a two-dimensional gel that was run to separate the proteins found in a snake toxin. Locate the prominent dark spot in the upper right-hand quadrant on the two-dimensional gel and answer these questions.  Name one thing that you can infer about the amino acid composition of this major spot from the isoelectric focusing direction of the gel?  What is one thing that you can infer from the PAGE direction of this gel about this protein? Image Source: Hatakeyama, D. M., de Morais-Zani, K., Serino-Silva, C., Grego, K. F., Sant’Anna, S. S., Fernandes, W., Aniz, P. A. E. A., Torquato, R. J. S., Tanaka, A. S., Sanz, L., Calvete, J. J., & Tanaka-Azevedo, A. M. (2018). Examination of biochemical and biological activities of Bothrops jararaca (Serpentes: Viperidae; Wied-Neuwied 1824) snake venom after up to 54 years of storage. Toxicon (141), 34–42. Image Description  A two-dimensional Electrophoresis (2-DE) gel showing the separation of proteins. The x-axis represents the pH range from 3 to 10, and the y-axis represents molecular weight from 15 kDa to 250 kDa. Multiple protein spots are visible, indicating their isoelectric points and molecular weights. The label A marks a significant area of interest on the gel. The gradient shading reflects the concentration of proteins, with darker spots representing higher protein abundance.

The Gly and Pro, as well as modified prolines that have a hy…

The Gly and Pro, as well as modified prolines that have a hydroxy group on the Pro ring or (Hyp), are the principal amino acids comprising collagen. Consider this excerpt from this article: Destabilization of osteogenesis imperfecta collagen-like model peptides correlates with the identity of the residue replacing glycineKonrad Beck, Virginia C. Chan, Nigel Shenoy, Alan Kirkpatrick, John A. M. Ramshaw, and Barbara BrodskyPNAS April 11, 2000 97 (8) 4273-4278; https://doi.org/10.1073/pnas.070050097 “Gly-Pro-Hyp is the most common, as well as the most stabilizing, triplet in collagen (5). The presence of Gly at every third residue is considered essential….” a. If there is a genetic mutation that interrupts this pattern of Gly at every third position, bones do not develop properly. They are characteristically brittle, resulting in a genetic disease known as osteogenesis imperfecta. Why are repeating Gly residues essential to the formation of a proper and strong collagen structure?  (1 pts.) b. Vitamin C is required for the biochemical reaction that puts the modifying OH group on a Pro in collagen. How does the presence of this group affect the relative overall strength of the collagen? (1 pts.)

The calcitonin gene can encode either the hormone calcitonin…

The calcitonin gene can encode either the hormone calcitonin or a protein called calcitonin-gene-related peptide depending on which 3′ cleavage site is used. In the thyroid gland, cleavage and polyadenylation occur after the fourth exon leading to calcitonin production. However, in the brain, the exact same transcript is cleaved after the sixth exon yielding calcitonin-gene-related peptide. This is possible because of a process known as:

Upload an image of your answers to this question. If you de…

Upload an image of your answers to this question. If you decided to use a PIPES buffer (piperazine-N,N’-bis(2-ethanesulfonic acid)) instead of the HEPES buffer used in this procedure, how many moles of the protonated form of PIPES and how many moles of deprotonated form of PIPES would you have to add to 1.0 liter of water to prepare a 0.10 M PIPES buffer at pH 7.5? The appropriate pKa is 6.76.  Show your work for full or partial credit. (Note: The sulfonic acid portion of the molecule is a strong acid and hence, fully deprotonated at pH 7.5.)