As described in Chapter 12, the collection of metabolic processes within a cell or organism is a complex interconnected set of pathways that include differing enzymes, cofactors, substrates, and products. Both the Braunschweig Enzyme Database (BRENDA) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) provide a means to retrieve information about metabolic reactions in different organisms. In this exercise, the BRENDA and KEGG databases will be searched for biochemical information about various enzymes and cofactors used in catabolism.

Instructions

1. Go to http://www.brenda-enzymes.org.

2. For Questions 1-5 and 10 from the assessment section, use Steps 3-4.

3. For this exercise, only the general search features of the BRENDA database are required; these options/functions are highlighted in the image below.

Entry Field

Search Options 4. Note the following options, shortcuts, and icons:

a. To narrow or refine a search, use the search options, such as “cofactor,” “enzyme, ligand,” and others.

b. To quickly navigate within an enzyme’s page, use the left menu. For example, click on:

i. Enzyme Nomenclature and choose a desired section (e.g., Pathway).

ii. Enzyme-Ligand Interactions and choose desired section (e.g., Cofactor).

iii. Functional Parameters and choose the desired parameter.

c. To view a reaction diagram, click on the beaker image in the “Reaction” and “Substrate” tables, which are in the “Enzyme Nomenclature” and “Enzyme-Ligand Interactions” menu options (left menu).

5. For Questions 6-9 from the assessment section, use the KEGG database:

http://www.kegg.jp.

6. Instructions for using the KEGG database are found in Bioinformatics Exercise 10.2.

7. Use these combined functions and databases to address the questions below.

Assessment Questions

1. As indicated in the BRENDA database, the classification number 2.7.1.1 refers to ________, and ATP is a _________ in the reaction catalyzed by this enzyme.

A. phosphoglucokinase; co-substrate

B. hexokinase; co-product

C. hexokinase; co-substrate

D. phosphoglucokinase; co-product

2. As indicated by the BRENDA database, the enzyme hexokinase is NOT used in which pathway?

A. sucrose synthesis

B. sucrose degradation

C. streptomycin synthesis

D. glycolysis

3. The enzymes classified as 1.1.1.1 and 1.1.1.2 are both oxidoreductases that catalyze reversible reactions. Which cofactors are used as co-substrates by these enzymes when catalyzing reduction reactions?

A. For 1.1.1.1 NADPH; For 1.1.1.2 NADH

B. For 1.1.1.1 NADH; For 1.1.1.2 NADPH

C. For 1.1.1.1 NADP + ; For 1.1.1.2 NAD +

D. For 1.1.1.1 NAD + ; For 1.1.1.2 NADP +

4. Which enzyme class does NOT use biotin as a cofactor?

A. Lyases

B. Ligases

C. Oxioreductases

D. Transferases 5. According to Table 14-1 and the BRENDA database, the oxoglutarate dehydrogenase enzyme that is specifically characterized as ___________ uses lipoic acid as a cofactor to catalyze _______________ reactions.

A. succinyl-transferring; alkyl transfer

B. succinyl-transferring; acyl transfer

C. NADP + -dependent; acyl transfer

D. NADP + -dependent; alkyl transfer

6. In the KEGG database, the pathway map for glycolysis/gluconeogenesis indicates that ___ unique enzyme/s can catalyze the conversion of α-D-glucose to α-D-glucose-6-phosphate (αD-glucose-6P).

A. 2

B. 3

C. 4

D. 1

7. How many steps are required to convert α-D-glucose-6-phosphate (α-D-glucose-6P) to glycerate-1,3-bisphosphate (glycerate-1,3P 2 )?

A. 6

B. 2

C. 4

D. 5

8. Among the steps highlighted in question 7, which enzyme catalyzes an irreversible reaction?

A. fructose-bisphosphate aldolase

B. 6-phosphofructokinase 1

C. glucose-6-phosphate isomerase

D. glyceraldehyde-3-phosphate dehydrogenase

9. As indicated in the KEGG pathway map, the reaction catalyzed by hexokinase (2.7.1.1) is irreversible; according to the BRENDA database, which metabolite inhibits this enzyme?

A. ATP

B. AMP

C. Citrate

D. All of these metabolites inhibit hexokinase