BCH 4053 PRE-TEST 4 GROUP NAME _____________________ July 24, 1996 This test is take-home and open book, and it is intended that all members of the group contribute to completing it. Only one copy is to be submitted by the group, and all members who participated should sign their names below. Test is due by 1:30 pm on Monday, July 29. __________________________ __________________________ __________________________ __________________________ __________________________ __________________________ Points (8) 1. Identify by name of the enzyme the reaction(s) of glycolysis which: (a) are isomerases (b) consume ATP as a substrate coenzyme (c) produce ATP as a substrate coenzyme (d) interconvert an aldose and a ketose (9) 2. For the following enzymes of glycolysis, give the structure of the non-coenzyme reactants and products, the name (or abbreviation) of coenzyme substrates or products, and indicate whether the reaction is normally near equilibrium or metabolically irreversible. (a) phosphofructokinase-1 (b) glyceraldehyde-3-phosphate dehydrogenase (c) enolase (12) 3. Parts of the glycolytic pathway can mediate the catabolism of other substances. For example, hydrolysis of triglycerides in adipose tissue produces not only free fatty acids, but glycerol. The glycerol cannot be re-activated in adipose tissue, but can be taken up from the blood by liver, and there converted to pyruvate. The initial reactions of its metabolism in liver are the following: glycerol + ATP ----> L-à-glycerol phosphate + ADP L-à-glycerol phosphate + NAD+ ----> dihydroxyacetone + NADH phosphate (a) Give the overall pathway by which glycerol is converted to pyruvate, showing the structures of the carbon-compound intermediates. (Substrate coenzymes may be abbreviated). (b) Starting with [sn 1-14C] glycerol, circle the radioactive carbon of each intermediate, showing which carbon of pyruvate will become labeled. (c) Give the overall stoichiometry of the reaction, showing the total production of ATP and NADH. (3) 4. A deficiency of the enzyme hexokinase in red cells affects the binding of oxygen to hemoglobin. Explain why, and predict whether oxygen dissociation (i.e. P50) would be higher (looser binding) or lower (tighter binding). (12) 5. Phosphorylase is an allosteric protein, existing in an R (active) and a T (inactive) conformation. The T/R ratio is affected by several allosteric "effectors", which activate or inhibit enzyme activity. For each of the following compounds, indicate whether it primarily affects the phosphorylated or non-phosphorylated form of the enzyme, whether it shifts the T/R ratio toward the T form or the R form, and whether the effect is an activation or inhibition of enzyme activity. Phosphorylated or Activation or Effector Non-phosphorylated T or R Inhibition ATP ______________ ____ ___________ AMP ______________ ____ ___________ Glucose ______________ ____ ___________ Glucose-6- ______________ ____ ___________ phosphate (6) 6. Draw structures of oxaloacetate to show the labeling pattern it would have from incorporation of 14C from [2-14C]acetyl-CoA after (a) one turn of the TCA cycle. (b) two turns of the TCA cycle. (c) three turns of the TCA cycle. (8) 7. Give the structure of the products formed when sedoheptulose-7- phosphate reacts with glyceraldehyde-3-phosphate in the presence of: (a) transaldolase (b) transketolase (20) 8. As an exercise in tracing radioactive labels, use the enzymes of the pentose phosphate pathway, as well as any enzymes you may need from the glycolytic pathway, to show how radioactivity from [2-14C]- glucose (i.e., glucose labeled with carbon-14 in position 2) can be converted to 14CO2. Show the structures of the intermediates of your pathway, circling or starring the radioactive atoms in each structure, and give the name of the enzyme catalyzing each step. (22) 9. In your first set of metabolic pathways, you have encountered many enzymes. Following is an alphabetical list of many of them: (1) aconitase (2) aldolase (3) citrate synthase (4) enolase (5) fructose-1,6-bisphosphatase (6) fructose-2,6-bisphosphatase (7) fumarase (8) galactose-1-phosphate uridylyltransferase (9) gluconolactonase (10) glucose-6-phosphatase (11) glucose-6-phosphate isomerase (12) glucose-6-phosphate dehydrogenase (13) glyceraldehyde-3-phosphate dehydrogenase (14) glycogen phosphorylase (15) glycogen synthetase (16) hexokinase (17) isocitrate dehydrogenase (NAD+ requiring) (18) isocitrate dehydrogenase (NADP+ requiring) (19) à-ketoglutarate dehydrogenase complex (20) malate dehydrogenase (21) malic enzyme (22) phosphoenolpyruvate carboxykinase (23) phosphofructokinase-1 (24) phosphofructokinase-2 (25) phosphoglucomutase (26) 6-phosphogluconate dehydrogenase (27) phosphoglycerate mutase (28) phosphoglycerate kinase (29) pyruvate carboxylase (30) pyruvate dehydrogenase complex (31) pyruvate kinase (32) ribulose-5-phosphate isomerase (33) ribulose-5-phosphate epimerase (34) succinate dehydrogenase complex (35) succinyl-CoA synthetase (36) transaldolase (37) transketolase (38) triose phosphate isomerase (39) UDP-glucose pyrophosphorylase (40) UDP-glucose 4'-epimerase Choose enzymes from the list that are described by the following statements, and place the number or numbers of the enzyme in the blank to the left of the statement. In many cases, more than one enzyme will apply. Given in parenthesis after the statement is a target number of enzymes for you to identify, though there may be more than that target number on the list. (a) __________________ CO2 is a substrate or a product. (four enzymes) (b) __________________ ATP is a substrate or a product. (four enzymes) (c) __________________ GTP is a substrate or a product. (two enzymes) (d) __________________ Coenzyme A is a substrate or a product. (three enzymes) (e) __________________ NADH is a substrate or a product. (three enzymes) (f) __________________ NADPH is a substrate or a product. (three enzymes) (g) __________________ Thiamine pyrophosphate is a prosthetic group. (three enzymes)