Purine Synthesis: Synthesis of Purine RiboNucleotides

In 1948 “John Buchanor” obtains the first clues as to how this process occurs. Denovo by feeding a variety of isotopically labeled compounds to pigeons and chemically determining the position of the labeled atoms in their excreted Uric acid.

The two purine nucleotides of nucleic acids are AMP and GMP containing the purine bases Adenine (A) and Guanine (G).

The results of the studies are Purine synthesis.

• What is Tautomerization and its changes? (Basics)
• Nucleic Acids are the Molecular Language of life
• Basic Components of Nucleic Acids – Purines and Pyrimidines

The biosynthetic organs of Purine ring atoms note that C4, C5, and N7 come from a single Glycine molecule but each of the other atoms is derived from an independent precursor.

• N1→ came from Asparagine
• C2 and C8 → came from formate (THF)
• N3 and N9 → came from Glutamine
• C4, C5, and N7 → came from HCO₃^–

Purine Synthesis Pathways

Purine synthesis can be explained in two different pathways

1. De-Novo Pathway
2. Salvage Pathway (also called Dust-bin Pathway)

De Novo Purine Synthesis

In this De novo synthesis of purines, each atom in the purine nucleotide came from different sources as mentioned above structure and data. There are 3 major steps are involved in this Purine synthesis pathway.

1. Ribose-5-Phosphate to IMP synthesis
2. Synthesis of AMP from IMP
3. Synthesis of GMP from IMP

Ribose-5-Phosphate to IMP synthesis

Step 1: Amination

The starting material for purine biosynthesis is Ribose-5-P, a product of the Hexose MonoPhosphate Shunt or Pentose Phosphate pathway (HMP Shunt). The ribose-5-P is converted into phosphoribosyl pyrophosphate by Pyrophospho Kinase in this reaction ATP is consumed.

Step 2: Addition of N9

One nitrogen is added on Ribose-5-P, to form 5-phosphoribosyl-1-amine (PRA). The nitrogen is donated by Glutamine. Ribose-5-Phosphate is derived from PRPP.

The reaction needs energy from ATP hydrolysis. This is the rate-limiting enzyme of this pathway. This step is inhibited by azaserine, the anticancer drug.

Step 3: Incorporation of C4, C5, and N7

The phosphoribosyl amine (PRA) is condensed with glycine it forms Glycinamide ribotide (GAR). This reaction is catalyzed by GAR Synthase.

Step 4: Adition Of C8

Glycinamide ribotide is converted into a Formyl glycine amide ribotide (FGAR). This reaction is catalyzed by transformylase. Here Formyl donor is N¹⁰-Formyl-THF.

Step 5: Addition of N3

Formyl Glycine ribotide is converted into Formylglycinaidine ribotide (FGAM) in the presence of the enzyme FGAM synthetase. Here amide donor is Glutamine and it is ATP consumed reaction. This enzyme is also inhibited by azaserine.

Step 6: Cyclisation (Closure of Ring)

FGAM is converted into 5-amino imidazole ribotide (AIR). This reaction is catalyzed by AIR Synthetase. Here ATP is consuming.

Step 7: Addition of C6

Amino imidazole ribotide is converted into 5-amino-4-Carboxy-Amino-Imidazole Ribonucleotide (CAIR). This reaction catalyzed by AIR carboxylase. In this reaction, Carbonic acid is substituted on a 4^th carbon atom as in the form of the Carboxyl group (CAIR).

This carbon dioxide fixation reaction does not require biotin or ATP.

Step 8: Addition of N1

Carboxy Amino Imidazole has converted into 5-AminoImidazole (N-Succinylocarboxamide) ribotide (SACAIR). This reaction is catalyzed by SACAIR synthetase. In this reaction, one Aspartic acid linked with Carboxyl group ATP is consumed.

Step 9: Removal of Fumaric acid

SACAIR is converted into 5-AminoImidazole-4-CarboxyAmide Ribotide (AICAR). This reaction is catalyzed by Adenosuccinate Lyase. The linked Aspartic acid hydrolyzed as Fumarate, which directly enters into TCA cycle.

The amino group of aspartic acid becomes the first nitrogen of the purine ring.

Step 10: Addition of C2

AICAR is converted into 5-FormaminoImidazole-4-Carboxamide Ribotide (FAICAR). This reaction is catalyzed by Transformylase. Here formyl group donor is N¹⁰-Formyl THF. This carbon is derived from the one-carbon pool. In folic acid deficiency, this step is blocked; hence orange-colored FAICAR is excreted in the urine.

Step 11: Cyclization

FAICAR is converted into Inosine Mono Phosphate (IMP) by the catalyzation process. This reaction is catalyzed by IMP Cyclohydrolase. It contains the purine, hypoxanthine.

Synthesis of AMP from IMP

IMP is the central intermediate of both AMP and UMP.

Step 1: IMP to AdenyloSuccinate

The IMP is converted into adenyloSuccinate by taking Aspartate and GTP, Which gives the power by the UTP to GTP and inorganic phosphate. This reaction is catalyzed by Adenylo Succinate synthatase.

Step 2: AdenyloSuccinate to AMP

AdenyloSuccinate is converted into AMP by releasing Aspartate as in the form of Fumarate. This reaction is catalyzed by Adenylo Succinate Lyase.

Synthesis of GMP from IMP

Step 1: IMP to XMP

IMP is converted into Xanthosine Monophosphate in the presence if the enzyme IMP-dehydrogenase. This is the dehydrogenation.

Step 2: XMP to GMP

XMP is converted into GMP by the enzyme GMP synthase. Here Amino group donor is Glutamate.

Regulation of Purine Biosynthesis

Activation:

• The activator molecule for Purine synthesis is PRPP, which activates the enzyme AmidoPhospho Ribosyl transferase
• The initiator molecule for the synthesis is Ribose-5-Phosphate. The optimum concentration of Ribos-5-Phosphate is maintained by the enzyme Glucose-6-Phosphate dehydrogenase, which is the regulatory enzyme of Hexose Mono Phosphate Shunt.

Inhibition:

• The rate-limiting enzyme “Ribose-5-Phosphate Pyrophospho kinase” the enzyme inhibited Nucleotides AMP, ADP, ATP, GMP, GDP, GTP by Feedback inhibition mechanism.
• The Enzyme Amino phosphoribosyl transferase is inhibited by AMP, ADP, ATP, GMP, GDP, GTP adenylosuccinate and XMP.
• The Amp inhibits Adenylo succinate synthatase GMP inhibits IMP dehydrogenase.

Salvage Pathway (or) Dust-bin Pathway

This is another type of Purine Nucleotide Synthesis from scratch. So this is also called the “Dust-bin pathway”. Most cells have an active turnover of many of their nucleic acids, results in Adenine, Guanine, and Hypoxanthine.

• Pyrimidine Synthesis Pathway: Synthesis of pyrimidine derivatives
• Pyrimidine Catabolism: UMP and CMP degradation Pathway
• Purine Catabolism and its Uric Acid formation
• Purine Synthesis: Synthesis of Purine RiboNucleotides

Purines that result from the normal turnover of cellular nucleic acids or that is obtained from the diet and not degraded. It can be reconverted into Nucleoside triphosphate and used by the body. This is referred to as the “Salvage pathway” for Purines.

Two enzymes are involved in this pathway.

APRT Enzyme

• APRT means Adenosyl Phosphoribosyl Transferase (APRTase)
• This enzyme catalyzes the reaction of Adenine to AMP conversion. In this reaction the secondary substrate is PRPP and byproduct is PPi.

Adenine + PRPP ↔ AMP + PPi

HGPRT Enzyme

• HGPRT means Hypoxanthine-Guanine Phospho Ribosyl Transferase (HGPRTase).
• This enzyme catalyzes the reaction of GMP formation from Hypoxanthine and PRPP.

Hypoxanthine + PRPP ↔ IMP + PPi

Guanine + PRPP ↔ GMP + PPi

The salvage pathway is of special importance in tissues like RBC and brain where the de-novo pathway is not operating.

The origin of the carbon and the nitrogen atoms of the purine ring system, determined by John Buchanan using an isotopic tracer experiment.

The detailed explanation of Purine biosynthesis is first explained by Buchanan and G.Robert Greenberg in the 1950s.

Multiple Choice Questions (MCQ)