BCH 361 Advanced Biochemistry I Fall 2020 Metabolic Pathway Project Discussion & Conclusion Guidelines This time you will be submitting the discussion and conclusion of your metabolic pathway. Presentation: Your work will be typed, double spaced (except for the references – they are single spaced). DO NOT INCLUDE A TITLE PAGE. Put a title and your name at the top. We will be strictly enforcing the following page limits: 3 pages for the text, 1 page for your diagram of the pathway, 1 page for the diagram of the mechanism, 2 pages for the references you referred to in the text. Any part of your work falling outside of these limits will not be graded. This is not an English essay. It’s more like a review article. Therefore, please break up your work with some subheadings. These should let the reader know what to expect in the following text, but should not be questions. Content: You will address parts 2d-g as indicated in the original metabolic pathways project description. Additional details on these points:    2d: you will provide a written description for the mechanism for ONE of the enzymes in your pathway that we have not already discussed in BCH 361. This enzyme should catalyze an isomerization, C-C bond cleavage or formation, or an unusual redox reaction. Remember to refer to your drawing in the text. 2e: when discussing the regulation of the pathway, don’t forget to refer to your pathway diagram, especially when discussing the logic of the regulation point(s). If you can’t find any sources for the regulation, make an educated guess based on the BCH361 lectures and the similarity of the pathway to pathways we have studied in class. 2g: you do not need any literature sources for this! Simply count up the number of ATP or ATP equivalents, NADH and FADH2, and then convert these to the maximum possible ATP for the organism based on the information in the BCH 361 lectures. You do not need to cite the BCH 361 lectures for the conversions. Don’t forget to cite your sources! Do not copy and paste from your sources. Instead, you will paraphrase – this does not mean rearranging their words or replacing occasional words. Even better, completely integrate the information from your sources into your own thoughts and words. Pathway Diagram: Include the pathway diagram from the introduction, along with the title and caption. Make any corrections suggested by the marker of the introduction. Indicate the regulation point(s) of the pathway along with the regulating agent(s). See the BCH361 lectures for examples of how to do this. Enzyme Mechanism Diagram: Draw the mechanism for the enzyme. Provide a title and caption for this. Don’t forget to cite your source(s). Make sure you use arrows in a chemically correct sense. Curved arrows need to show the direction of electron flow. 1 BCH 361 Advanced Biochemistry I Fall 2020 Personal Perspective: Own this assignment! If you have a particular interest that this pathway relates to, try to work this into your presentation. Not only will you enjoy the project more, but your reader will enjoy your work more. References: The reference section will be single spaced. You will not include any annotations for your references this time. Include only the references you cite in your introduction. It is not absolutely necessary to use the ACS Biochemistry style, but using it will be easier for the markers. Use the same style throughout your work; this is what is meant by consistency. Make sure that you have acknowledged your sources; not doing so is a violation of Policy 60. 2 BCH 361 Advanced Biochemistry I Fall 2020 Rubric for Pathway Introduction Presentation Description Grade Your project is well presented: You have included a title and your name. The type face is of an appropriate style and size. Your text is double spaced. Your references are single spaced. Your pages are numbered. Your work falls within the page limits. 2 Your presentation is somewhat problematic. There are 1-2 problems such as a title and your name missing, page numbering, font style and size, line spacing the inclusion of a title page, exceeding the page limits. 1 Your presentation has more than 2 problems with the title, name, page numbering, font style and size, line spacing, or exceeding the page limits. 0 Writing Style Description Grade The writing style is consistent with an academic review paper. Subheadings are used to break the text into logical segments. The writer shows a good command of English. The text flows well. Literature information has been thoroughly integrated into the writer’s thoughts. 5 The writing style may be too casual for an academic review paper or subheadings have not been used in a logical manner. The writer shows a good command of English. The text flows well. Literature information has been thoroughly integrated into the writer’s thoughts. 4 The writing style and subheadings are not appropriate for an academic review paper. Literature information has not been well integrated into the writer’s thoughts; it is paraphrased. The writer otherwise shows a good command of English and the text flows well. 3 The writing style and subheadings are not appropriate for an academic review paper. Literature information has not been well paraphrased. The writer may show a poor command of English or the text may not flow well. 2 The writing style and subheadings are not appropriate. Literature information has not been well paraphrased. The writer is having difficulty with English but although the text does not flow well, the ideas are presented in a logical order. 1 There are major problems with the writing style and use of English. The ideas do not flow well. Literature information may be quoted instead of paraphrased or integrated into the text. 0 3 BCH 361 Advanced Biochemistry I Fall 2020 Content Description Grade Your text identifies the pathway. You have clearly identified the enzyme whose mechanism you describe. You have clearly identified similar mechanisms, if any, in the BCH 361 lectures. You have provided a logical explanation for this similarity. You have clearly identified the regulatory points of the pathway, the agents that regulate the pathway and how this regulation is achieved (the regulatory mechanism). You have a concise explanation for how this regulation is logical. You have accurately and concisely accounted for the energy production or use by your pathway and converted this to ATP equivalents as appropriate for the organisms that use the pathway. Your text refers to your diagrams. 15 Your text clearly identifies the pathway. You have identified the enzyme whose mechanism you describe, and the regulatory point(s) and agent(s). You have described the mechanism of the enzyme and the way the pathway is regulated. You have accurately accounted for the energy production or use by the pathway and converted this to ATP equivalents. However, you have a problem with one of: identifying similar enzymatic mechanisms, justifying the similarity, complete and concise explanation for the regulation, or have not referred to your diagrams. 12 Your text clearly identifies the pathway, the enzyme whose mechanism you show, and the regulatory methods used on the pathway. You have accurately accounted for the energy production or use by the pathway. There are minor problems with correctly identifying similar enzymatic mechanisms, providing logical explanations, or converting the energy of the pathway to ATP equivalents appropriate for the organisms using the pathway. 9 Your text clearly identifies the pathway and the enzyme whose mechanism you show. You may have significant problems with describing the mechanism and providing a logical explanation for a similar mechanistic strategy; or describing the regulatory methods and logic for the pathway. 6 The text identifies the pathway. There is no description of how the pathway is 3 regulated. There may be other significant problems with the content of your work. The text, if present, fails to identify the pathway, the regulatory points and the energy production or use by the pathway. 0 4 BCH 361 Advanced Biochemistry I Fall 2020 Pathway Diagram Description Grade Your pathway is clearly drawn with properly drawn structures of all the substrates and products of the reactions (except things like ATP and NAD). You have clearly identified the regulatory point(s) along with the regulatory agent(s). You have named each of the enzymes and the metabolic intermediates. You have included a title and caption and cited your source(s). 3 The pathway has some problems with clarity, the drawing of structures, indicating the regulatory agents, naming enzymes or intermediates, the title or caption. Regulatory points are clearly indicated. 2 The pathway has some problems with clarity, the drawing of structures, naming enzymes or intermediates, the title or caption The pathway does not show the regulatory points. 1 The pathway is poorly drawn or missing. There is no title or caption. 0 Enzyme Mechanism Diagram Description Grade The mechanism is clearly drawn with properly drawn structures of the substrate, intermediates and product. Arrows are used correctly to show the movement of electrons, equilibrium, or resonance. The mechanism makes chemical sense. You have included a title and caption and cited your source(s). 4 The mechanism has problems with clarity or the drawing of the structures. Arrows are used correctly. A title and caption are included, and your source(s) are cited. 3 The mechanism is poorly drawn (eg. incorrect use of arrows) or the title and caption are missing. 2 The mechanism is poorly drawn; it does not make chemical sense. There is no title or caption. 1 There is no mechanism diagram. 0 5 BCH 361 Advanced Biochemistry I Fall 2020 Originality, Insight and Personal Perspective Description Grade You have demonstrated your interest in the pathway. Your presentation, while academic, shows your interests, imagination, and/or personality. You regularly make logical inferences rather than relying on sources outside of course material where they are not needed. 3 Your work shows an interest in the pathway and shows some of you imagination and personality. You have avoided using sources outside of the course material when you could make inferences. 2 It is difficult to discern your interest in the pathway. Your presentation shows some imagination. You have a reliance on sources outside of the course materials instead of making inferences. 1 Your project does not demonstrate any particular interest in the pathway or imagination. You rely on sources outside of the course materials. 0 Citation and Reference Style and Formatting Description Grade The references and citations are provided in an appropriate style, eg. ACS Biochemistry. The reference and citation style is consistent. The references are all cited. The literature is cited appropriately (you have acknowledged your sources). Most of the sources are primary literature articles. 3 The references and citations are provided in an appropriate style. The reference and citation style is consistent. The references are all cited. The literature is cited appropriately. Some of the sources are primary literature sources. 2 The references and citations are in an appropriate style, but lack consistency. There may be minor problems with acknowledging literature sources. Many of the sources are not primary literature. 1 The references and citations are not in an appropriate style (eg APA or MLA). There are major problems with acknowledging your sources. 0 Deadline and late penalties: Due date: 11:59 pm, 4 December 2020 A 7 mark deduction if submitted within the first 24 hours after the deadline A 14 mark deduction if submitted between 24 and 48 hours after the deadline. A grade of 0 for work submitted more than 48 hours after the deadline. 6 BCH 361 Advanced Biochemistry I Fall 2020 Metabolic Pathways Project The goal of this project is to apply the concepts and ideas from the lectures of BCH 361 to other related metabolic pathways. This project will be submitted in parts to Turnitin via D2L. 1. Select a topic from this list. Sign up for the topic using the project group sign up on D2L. Each is limited to 50 students. Sign up early for the best chance of getting your first choice. a. Topic 1: Entner-Douderoff Pathway b. Topic 2: L-Ascorbate Biosynthesis c. Topic 3: D-Tagatose Degradation d. Topic 4: L-Rhamnose Degradation e. Topic 5: L-Arabinose Degradation f. Topic 7: meso-Tartaric Acid Metabolism g. Topic 8: Glyoxylate Cycle 2. Research your topic. You don’t need to do all of this at once. Plan your work around the submission deadlines and pace yourself so you don’t try to do it all at the end. a. What is the pathway? (Sometimes there are multiple pathways; choose the most appropriate one(s)). You need to show full structures for each of the substrates and identify the enzyme that catalyzes each reaction. Is this pathway similar to any other pathway we have discussed in the course? b. What organisms use the pathway? c. What is the purpose or importance of the pathway? d. How do the enzymes work (what are the mechanisms)? How do these compare to mechanisms of other enzymes discussed in the lectures? e. How is the pathway regulated? (This may not be known; make inferences and back up your position) f. Does the pathway feed into glycolysis, or the TCA cycle, or some other pathway? g. How much energy is generated (or used) in the pathway in terms of molecules such as ATP, NAD(P), and FAD? 3. Prepare and submit an annotated bibliography. a. Identify at least three journal articles that can serve as sources for your project. For three of these articles, outline in no more than three sentences per article the information it will provide towards your project. This does not need to be your entire reference list! b. Page limit = ½ page. c. SUBMISSION DEADLINE: 2 October 2020 4. Prepare and submit your Introduction. a. This report will include parts a-c in the above Research section. b. Include the pathway (each has at least two steps) showing the structures of the metabolites and naming each enzyme. Draw this out yourself; do not copy and paste from the literature or websites, but do cite your source(s). c. Include your reference list of works cited in the outline and introduction. d. Page limit = 2 pages e. SUBMISSION DEADLINE: 30 October 2020 [Type here] 1 BCH 361 Advanced Biochemistry I Fall 2020 5. Prepare and submit your final report. a. This report will focus on parts d-g in the above Research section. Include your Introduction (parts a-c). b. Draw the mechanism for at least one of the reactions that is not a simple phosphorylation/dephosphorylation. If possible, indicate the active site residues that are involved in the mechanism. c. You may wish to divide your report into sections as is normally done in writing review articles. d. Include your reference list with all the works cited. e. Page limit = 6 pages including Introduction f. SUBMISSION DEADLINE: 4 December 2020 Notes: 1. Submission Guidelines. Prepare your work as a PDF or DOC file for submission to Turnitin on D2L. Work exceeding the page limits will not be graded. Please be aware of the different submission deadlines and plan your work accordingly. Late work will be penalized. Early submissions are encouraged. Formatting: Your work will be double-spaced and have 1 inch (2.5 cm) margins, and use a standard 11 or 12 point business font. The reference section will be single spaced for each reference, and double spaced between references. If your work has more than one page, number the pages. Make sure your name is on the upper right corner of the first page of your work. Drawing structures and pathways: You may draw the structures by hand and photograph or scan them, or use software such as ChemDraw. Pyranose forms of sugars should be in a chair, boat, or twist-boat conformation. Do not show Haworth projections. 2. Originality and Academic Integrity. You are encouraged to be original in your presentation with regards to the perspective you take. However, you must acknowledge the work of others (your sources) using a standard format. Be consistent. A good format for your references and citations would be to use the ACS Biochemistry format (like in the lab), but to add in the title of the journal article. Do not quote your sources unless you want to comment on their wording. Instead, paraphrase. Student Learning Support offers a number of workshops, some of which deal with paraphrasing and citations. Failure to acknowledge your sources could result in an investigation for academic misconduct as outlined in Policy 60. If you are unsure what to do, or realize that you made an error, contact Dr. Johnson ASAP. 3. Grading Guidance on how each of these will be graded will be provided closer to the submission deadlines. [Type here] 2 BCH 361 Advanced Biochemistry I D-Tagatose Degradation Wendy Banh Van der Heiden, E., Delmarcelle, M., Lebrun, S., Freichels, R., Brans, A., Vastenavond, C.M., Galleni, M., and Joris, B. (2013). A pathway closely related to the D-tagatose pathway of gramnegative Enterobacteria identified in the gram-positive bacterium Bacillus licheniformis. Appl Environ Microbiol 79, 3511-3515. This paper presents the catabolism of D-tagatose in different bacteria. This degradation involves the use of different enzymes such as phosphorylase, dehydrogenase, kinase, and aldolase. The resulting products are dehydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (G3P). Bissett, D.L., and Anderson, R.L. (1974). Genetic evidence for the physiological significance of the D-tagatose 6-phosphate pathway of lactose and D-galactose degradation in Staphylococcus aureus. J Bacteriol 119, 698-704. This paper presents the microorganisms which initiate D-tagatose degradation. In Staphylococcus aureus, a gram-positive bacterium, the structural genes for the pathway are cotranscribed with specific genes that code for transport, phosphorylation, and cleavage. Donner, T.W., Magder, L.S., and Zarbalian, K. (2010). Dietary supplementation with D-tagatose in subjects with type 2 diabetes leads to weight loss and raises high-density lipoprotein cholesterol. Nutr Res 30, 801-806. In this paper, the researchers intended to determine the effect of oral D-tagatose in attenuating the rise in plasma glucose in subjects with type 2 diabetes mellitus. The results showed a potential use of D-tagatose among diabetic patients in terms of significantly lowering high-density lipoprotein cholesterol. Saunders, J.P., Donner, T.W., Sadler, J.H., Levin, G.V., and Makris, N.G. (1999). Effects of acute and repeated oral doses of D-tagatose on plasma uric acid in normal and diabetic humans. Regul Toxicol Pharmacol 29, 57-65. In this paper, the researchers intended to determine the effect of oral D-tagatose in increasing plasma uric acid. The subjects for the study were normal individuals and those which have type 2 diabetes. The results showed that D-tagatose is an effective sweetener with no adverse clinical effects. Buemann, B., Toubro, S., and Astrup, A. (1998). D-tagatose, a stereoisomer of D-fructose, increases hydrogen production in humans without affecting 24-hour energy expenditure or respiratory exchange ratio. The Journal of Nutrition 128, 1481-1486. This paper presents 24-hour energy expenditure among normal weight humans during consumption of D-tagatose or D-sucrose. D-tagatose Degradation Pathway 1 Wendy Banh D-Tagatose Degradation Pathway D-tagatose is a keto-hexose, which is a D-fructose's Carbon-4 epimer and a lower calorie natural-sugar found in milk and fruit products in small quantities (Van der Heiden et al., 2013). D-tagatose can replace sucrose in foods because their texture, taste, and cooking properties are the same. Due to its antihyperglycemic effect and lower glycemic index, D-tagatose seems to make a perfect sweetener. D-tagatose degradation pathway starts with transport across the organism's cell membrane and its phosphorylation to tagatose 1-phosphate. According to Van der Heiden et al (2013), tag 1-phosphate is further phosphorylated by tag 1-phosphate kinase enzyme to tagatose 1, 6-bisphosphate, which is finally broken down by gatYZ tagatose aldolase into dihydroxyacetone phosphate (DHAP) and Glyceraldehyde 3-phosphate (G3P) as illustrated below. D-tagatose Degradation Pathway Figure 1. Degradation pathway of D-Tagatose in Klebsiella Oxytoca. D-tagatose is used as a carbohydrate source to be transported and phosphorylated to yield the final products dihydroxyacetone phosphate and glyceraldehyde-3-phosphate (Van der Heidan et al., 2013). 2 D-tagatose Degradation Pathway 3 D-tagatose pathway is similar to the galactitol and lactose degradation pathway in that they all lead to the production of tagatose 1, 6-bisphosphate, which is found in the D-tagatose breakdown pathway. Van der Heiden et al. (2013) asserts that, Tagatose 1, 6-bisphosphate is cleaved to dihydroxyacetone phosphate (DHAP) and glyceraldehyde 3-phosphate (G3P). Organisms Utilizing D-Tagatose Degradation Pathway According to Van der Heidan et al. (2013), organisms using this D-tagatose pathway are enteric gram-negative bacteria, including Klebsiella pneumoniae, Salmonella enterica, and Klebsiella oxytoca. Enteric refers to the occurrences of these bacterium in the intestines. These bacteria break down the D-tagatose via the "PTS-dependent D-tagatose pathway encoded by tagKTH and gatYZ genes. This pathway has a few similarities to the glycolysis pathway. Both pathways catabolize carbohydrates into smaller units; Glycolysis uses glucose whereas Dtagatose degradation uses D-Tagatose. They both yield dihydroxyacetone phosphate (DHAP) and Glyceraldehyde-3-phosphate (G3P) during their cycle. However, in glycolysis DHAP and G3P are yield as intermediates to be used for the progression of the cycle, where D-tagatose degradation they are the final products. Importance of D-Tagatose Degradation Pathway In their pilot study, Donner et al. (2010) found that daily ingestion of D-tagatose with food by type 2 diabetic patients led to improvements in weight loss and increase of high-density lipoprotein (HDL) cholesterol. During this study, the rise of HDL cholesterol was not proportional to the observed weight loss, and there were no side effects or additional biochemical parameters observed on this glycemic control. In the confirmation of beneficial D-tagatose D-tagatose Degradation Pathway 4 effects on HDL cholesterol and weight loss on diabetic type 2 patients, D-tagatose treatment could reduce cardiovascular infections in this population at the highest risk. According to Saunders et al. (1999), D-tagatose, which is a D-fructose stereoisomer, is a ketohexose that is naturally occurring and is proposed to be used as a lower-calorie sweetener. This is because ingested amounts of D-tagatose are insufficiently absorbed, while those taken up are metabolized by a pathway comparable to the one for D-fructose in the liver. A study conducted by Saunders et al. (1999) that was aimed at determining whether repeated or acute oral D-tagatose doses would elevate uric acid in plasma in healthy humans and diabetes type 2 patients. Results of the study showed that, D-tagatose is a promising sweetener with no observed clinical adverse effects. Buemann et al. (1998) confirms that, D-tagatose increases the production of hydrogen in humans with no effect on energy expenditure for 24 hours or respiratory exchange proportion. Based on this study, the previous propositions that ketohexose D-tagatose would have a distinct thermogenic effect explaining its apparent absence of the net energy could not be confirmed. Though an increase in H2 production displays that a considerable amount of sugar is poorly absorbed. Therefore, D-tagatose can be used as an ingredient in ready to consume breakfast cereals, a variety of confectionaries, soft drinks, meals-replacement drink powder or mix, formula diets used for replacement of meals, and baked goods. D-tagatose Degradation Pathway 5 References Buemann, B., Toubro, S., and Astrup, A. (1998). D-tagatose, a stereoisomer of D-fructose, increases hydrogen production in humans without affecting 24-hour energy expenditure or respiratory exchange ratio. The Journal of Nutrition 128, 1481-1486. Donner, T.W., Magder, L.S., and Zarbalian, K. (2010). Dietary supplementation with D-tagatose in subjects with type 2 diabetes leads to weight loss and raises high-density lipoprotein cholesterol. Nutr Res 30, 801-806. Saunders, J.P., Donner, T.W., Sadler, J.H., Levin, G.V., and Makris, N.G. (1999). Effects of acute and repeated oral doses of D-tagatose on plasma uric acid in normal and diabetic humans. Regul Toxicol Pharmacol 29, 57-65. Van der Heiden, E., Delmarcelle, M., Lebrun, S., Freichels, R., Brans, A., Vastenavond, C.M., Galleni, M., and Joris, B. (2013). A pathway is closely related to the D-tagatose pathway of gram-negative Enterobacteria identified in the gram-positive bacterium Bacillus licheniformis. Applied Environmental Microbiology 79, 3511-3515.
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