Under aerobic conditions, bacteria degrade glucose via glycolysis. The most comm
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Question
Under aerobic conditions, bacteria degrade glucose via glycolysis. The most common form of glycolysis that you have studied in class is also called the Embden-Meyerhof-Parnas (EMP) pathway. Figure 1 shows few steps of the EMP pathway.
Figure 1: Simplified diagram of the glycolytic (EMP) pathway. In this figure, arrows indicate the carbon #1 and 6 in glucose. DHAP and G-3-P stand for dihydroxyacetone phosphate and glyceraldehyde-3-phosphate respectively. The phosphate groups are labeled Ph. Note that carbon numbers in G3P and the subsequent molecules in the pathway are not indicated since they will differ depending on the origin of G 3-P (directly from Y or from DHAP).
1) Two molecules are labeled X and Y. What are their actual names? (4 pts)
X:
Y:
2) In bacteria, other pathways contribute to the oxidation of glucose molecules. These pathways include the hexose monophosphate pathway (HMP, Figure 2A) and in some gram-negative bacteria the Entner-Doudoroff (ED) pathway (Figure 3). Gram positive bacteria do not metabolize glucose through the ED pathway. The second step of the HMP cycle is a decarboxylation reaction leading to the conversion of 6-phosphogluconate into ribulose-5- phosphate (Figure 2B).
Some bacteria are lacking the enzyme that convert product Y (see figure 1) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde 3-phosphate (G-3-P) and don’t have a functional ED pathway. Could these bacteria rely on the HMP pathway for the production of ATP? Briefly explain. (6 pts)
Figure 2: Oxidative HMP cycle. Panel A: Glucose is degraded into glyceraldehyde-3-phosphate and CO2 in a reaction cycle. Panel B: reaction leading to the release of CO2.
Figure 3: ED pathway: Glucose is converted in glucose-6-phosphate (not shown) and then glucose-6-phosphate is broken down into pyruvate via a phosphogluconate intermediate.
3) To determine which glucose oxidizing pathway species of bacteria use, scientists have design respirometric experiments using glucose labeled with radioactive carbon (14C) in various position. Then, scientists collect the CO2 released soon after the bacteria were fed with the radiolabeled glucose. It is assumed that the released CO2 is the result of the decarboxylation of BIOS S10 3 pyruvate (EMP and ED pathways) or 6-phosphogluconate (HMP pathway). Remember that, under aerobic conditions, the carboxyl group of pyruvate is released as CO2 when pyruvate reacts with coenzyme A to produce acetyl-coenzyme A.
a) Three separate analyses of glucose degradation by the gram-negative bacteria E. coli through radiorespirometry were performed using each time a different types of radiolabeled glucose molecules: glucose with carbon #1 labeled, or glucose with carbon #2 labeled, or glucose with carbons #3 and 4 labeled. The graph below (Figure 4) shows the results of the 3 experiments. Based on the pathways given in figures 1 to 3, rank (when possible) metabolic pathway (EMP, HMP, or ED) by order of predominance in E. coli? Briefly explain. (8 pts)
Figure 4: Graph showing the amount of 14CO2 release when glucose labeled on carbon #1 (14C(#1)), #2 (14C(#2)) and #3 AND 4 (14C(#3&4) were used to feed E. coli. The Y-axis represents the amount of 14CO2 released.
b) Based on your knowledge of the fate of acetyl-coenzyme A under aerobic conditions, which metabolic pathway is likely at the origin of the 14CO2 released at the later stage of the experiment when glucose is labeled on carbon #2 or on carbon #1? Based on your answer in part b), would you modify your answer from part a)? Explain your reasoning. (6 pts)
c) The following two graphs show the same types of respirometric experiments with two strains of bacteria (A and B). Based on this graph, identify which strain is gram-negative. Briefly explain. (6 pts)
Figure 5: Graph showing the amount of 14CO2 release at the early stage when glucose labeled on carbon #1 (14C(1)), #3 (14C(3)) and #4 (14C(4) were used to feed two different bacterial strains. The Y-axis represents the amount of 14CO2 released.
#1 Glucose Glucose-6-Phosphate CH2 OH CH20 Ph H-C-OH H-C-OH HO-C-H HO-C-H H-C-OH H-C-OH H-C-OH H-C-OH H-C-OH H-C-OH CH2OH CH20-Ph CH20-Ph CH20-Ph #1 CH20-Ph CH2 OH DHAP CH20-Ph G-3-P + Pi COO-Ph CH20-Ph Pyruvate C=OExplanation / Answer
1. X is Fructose-6-Phosphate
Y is Fructose-1,6-Biphosphate
2. Yes they can rely on the HMP pathway. The Glucose-6-phosphate can enter HMP nad is first converted into 3,6-Phosphogluconate followed by 3-Ribulose-5-Phosphate. The Ribulose-5-Phosphate is then converted into G-3-P which can re-enter the EMP pathway to form pyruvate and furtheron to form ATP.
3.a. Since E.Coli is a gram negative bacteria, ED pathway is predominant. Followed by the EMP pathway, since it can directly form G-3-P, it does not require the HMP pathway.
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