Exam #3

Question 1

List differences in the biosynthetic pathways of purines and pyrimidines.

Answer 1

Purines: (adenosine and guanosine)

• Gives the intermediate inosine monophosphate (IMP)
• Start with alpha-D-5-phosphate (R5P)
• nitrogenous base is built first on ribose.
• Activated intermediate 5-phosphoribosul alpha-pyrophosphate (PRPP) is a key metabolic regulator.
• Cross-talk regulation between ATP and GTP

Pyrimidines: (cytosine and thymine)

• Synthesized from uracil monophosphate (UMP); can be converted to CMP
• Nitrogenous ring is assembled first and attached to the base
• RNA nucleotides are built before modifying to get DNA nucleotides

Question 2

Explain key regulatory molecules and mechanisms of purine biosynthesis.

Answer 2

1. Cys 225 blocks the binding site of 2’-hydroxy nucleotide
2. ATP binding to the active site of ribonucleotide reductase –> conformation change occurs, the enzyme is active, reconnects the radical synthesis pathway and the radical storage pathway, and able to transfer the tyrosine radical to the alpha-site.
3. dATP inhibits the enzyme by causing a conformational change that separates the radical generation site and the radical storage site.

Question 3

Define the term “de novo”

Answer 3

“anew”

Question 4

Define the term “salvage” in terms of nucleotide biosynthesis.

Answer 4

means that nucleotides were made from the breakdown of nucleic acids. This is done using the PRPP intermediate. Beneficial because the ring doesn’t have to be rebuilt every time.

Question 5

Describe the unique aspects of DNA nucleotides

Answer 5

Its nucleotides have 2’-deoxyribose residues instead of the ribose residues

Question 6

Explain the regulatory mechanisms of ribonucleotide reductase.

Answer 6

• Enzyme would become inhibited if a substrate was brought in before the disulfide bond could be reduced, so the enzyme prevents this by having cysteine (disulfide form) block the active site. Only reduced will bind.
• ATP causes conformation change that inhibits enzyme by physically separating the radical synthesis pathway and the radical storage pathway
• dATP causes conformation that activates the enzyme and connects the two pathways

Question 7

5-Phosphoribosyl pyrophosphate (PRPP) is a key intermediate in nucleotide biosynthesis. Identify the bond that PRPP allows to be formed.

Answer 7

Allows the amine to be hydrolyzed off of glutamine and given to the PRA. Allows the nitrogenous base ring to form.

Question 8

Why does it make sense that AMP and GMP to regulate the synthesis of each other?

Answer 8

• Both come from IMP
• AMP requires Aspartate and GTP to make adenylsuccinate which then makes AMP.
• GMP requires ATP and H2O to hydrolyze glutamine to make GMP

Question 9

List two major differences between nucleotides used for DNA compared to those used in RNA.

Answer 9

• DNA uses 2’ deoxy nucleotides

- DNA uses thymidine instead of uridine.

Question 10

Ribonucleotide reductase has two subunits: alpha and beta. Briefly describe the overall reaction catalyzed by the enzyme and the biochemical role of each subunit.

Answer 10

Catalyzes the formation of 2’-deoxynucleotides (which makes DNA only)

• Alpha subunit: where reduction/chemistry takes place. Radicals are brought to alpha when ready. Catalytic is found on in this.
• Beta subunit: di-iron center. Contains tyrosine 122 radical. Very stable. Beta-enzyme is able to stabilize the tyrosine residue, makes it non-reactive. Generates and stores the radical required for the reaction.

Question 11

Ribonucleotide reductase uses a stabilized _______ intermediate in its reaction cycle.

Answer 11

radical.