Purine Synthesis: Synthesis of Purine RiboNucleotides

Purines are vital organic compounds that play a fundamental role in various biological processes, such as the synthesis of DNA and RNA, energy metabolism, and cell signaling. These compounds are essential building blocks in the production of nucleotides, which are the fundamental units of genetic material.

Purine biosynthesis is a highly regulated process responsible for synthesizing purines from simple precursors, such as amino acids and formate.

The production of these precursors requires a great deal of energy, so regulating purine biosynthesis is critical to ensure that cells only produce the necessary amounts of purines at the right time.

Additionally, purine biosynthesis is essential in producing essential signaling molecules such as cyclic AMP and GTP, which play key roles in cellular metabolism and signaling.

Understanding purine biosynthesis and the regulation of purine metabolism is critical for maintaining the body’s purine balance and preventing disorders such as gout, Lesch-Nyhan syndrome, and various immunodeficiency disorders. Therefore, research on purine biosynthesis is of significant importance for basic research and clinical medicine.

In 1948, “John Buchanor” obtained the first clues about 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.

AMP and GMP are nucleic acid purine nucleotides that contain the purine bases Adenine (A) and Guanine (G). The results of the studies concern purine synthesis.

What is purine synthesis?

Purine synthesis is the process by which purine nucleotides, such as adenosine and guanosine, are synthesized de novo within the body.

• Purine nucleotides are essential building blocks for DNA and RNA and critical components of various metabolic pathways and signaling molecules.
• The synthesis of purine nucleotides occurs through a series of enzymatic reactions that convert simpler precursors into final products, including converting amino acids and other metabolites into nucleotide bases.
• Purine synthesis is a tightly regulated process essential for maintaining the proper balance of nucleotides in the body and is crucial for various biological processes such as DNA replication, cell division, and energy metabolism.
• Dysregulation of purine synthesis can lead to multiple disorders, including gout and Lesch-Nyhan syndrome.

Sources of atoms in Purine Ring

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 biosynthesis pathway can be explained in two different ways

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 mentioned in the structure and data. There are three significant steps involved in this purine synthesis pathway. There are 10 steps of purine synthesis. Let us take the purine synthesis steps one by one.

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

Here are the steps of de novo synthesis of purine nucleotides.

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). In this reaction, ribose-5-phosphate is converted into phosphoribosyl pyrophosphate by pyrophosphokinase, and ATP is consumed.

PRPP + Glutamine + H₂O → PRA + Glutamate + PPi

(PRPP = 5-phosphoribosyl-1-pyrophosphate, PRA = 5-phosphoribosyl-1-amine)

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 for this pathway. This step is inhibited by azaserine, the anticancer drug.

Step 3: Incorporation of C4, C5, and N7

When phosphoribosyl amine (PRA) is condensed with glycine, glycinamide ribotide (GAR) is formed. This reaction is catalyzed by GAR synthase.

PRA + ATP → GAR + PPi

Step 4: Addition of C8

Formyl glycine amide ribotide (FGAR) is formed from glycinamide ribotide. This reaction is catalyzed by transformylase. The Formyl donor in this case is N10-Formyl-THF.

GAR + FGAM → FGAR + AICAR

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.

FGAR + ATP → FGAM + ADP

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 consumed.

FGAM → AIR + NH3

Step 7: Addition of C6

Amino imidazole ribotide is converted into 5-amino-4-carboxy-amino-imidazole ribonucleotide (CAIR). This reaction is catalyzed by AIR carboxylase. In this reaction, carbonic acid is substituted with a 4th carbon atom in the form of the carboxyl group (CAIR).

AIR → CAIR + H₂O

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.

CAIR → SAICAR + CO₂

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 is hydrolyzed into fumarate, which directly enters the 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-Formamino-Imidazole-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-coloured 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 purine, hypoxanthine.

SAICAR → IMP + Fumarate

Donor atom	Added features	Special	Product
Glutamine	N9	Rate-limiting	PRA
Glycine	C4, C5 and N7	ATP required	GAR
Methenyl-THFA	C8	–	FGAR
Glutamine	N3	ATP required	FGAM
–	Ring closure	ATP need	AIR
Carbon dioxide	C6	–	ACAIR
Aspartic acid	N1	ATP required	SAICAR
–	–	Fumarate removed	AICAR
Formyl THFA	C2	–	FAICAR
–	–	Ring closure	IMP

Synthesis of AMP and GMP from IMP

Adenosine monophosphate (AMP) and guanosine monophosphate (GMP) are essential nucleotides that play a crucial role in various biological processes.

• AMP and GMP are synthesized from the precursor inosine monophosphate (IMP) via different pathways.
• The synthesis of AMP from IMP is carried out through a series of reactions involving adding an amino group. In contrast, the synthesis of GMP from IMP consists of adding a formyl group.
• Understanding the mechanisms behind synthesizing these nucleotides is essential for gaining insight into their biological functions and the potential therapeutic applications of targeting their biosynthesis.

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 of the UTP to GTP and inorganic phosphate. This reaction is catalyzed by adenyl-succinate synthetase.

Step 2: AdenyloSuccinate to AMP

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

Synthesis of GMP from IMP

Step 1: IMP to XMP

In the presence of the enzyme IMP-dehydrogenase, IMP is converted into Xanthosine Monophosphate. This is called dehydrogenation.

Step 2: XMP to GMP

XMP is converted into GMP by the enzyme GMP synthase. Here, the amino group donor is glutamate.

Overall equation of Purine Synthesis

PRPP + Glutamine + ATP → IMP + PPⁱ + Glutamate + Fumarate + CO₂ + NH₃ + 2ADP

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” inhibited the nucleotides AMP, ADP, ATP, GMP, GDP, and GTP by a 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 of purine synthesis

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, resulting in adenine, guanine, and Hypoxanthine.

Purines that result from the normal turnover of cellular nucleic acids or that are obtained from the diet are 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 the brain, where the de-novo pathway is not operating.

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

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

Disorders of Purine Metabolism

Here’s the content for the “Disorders of Purine Metabolism” section in a point-wise format:

• Disorders of purine metabolism refer to a group of inherited metabolic disorders that affect the body’s ability to process purines.
• Gout is a common purine metabolism disorder that results from the deposition of uric acid crystals in joints and other tissues, causing inflammation and pain.
• Lesch-Nyhan syndrome is a rare disorder that affects the metabolism of purines and results in hyperuricemia, neurological symptoms, and self-injurious behaviour.
• Other disorders of purine metabolism include adenine phosphoribosyltransferase deficiency, purine nucleoside phosphorylase deficiency, and hypoxanthine-guanine phosphoribosyltransferase deficiency (HGPRT).
• These disorders can lead to various symptoms, including joint pain, muscle weakness, intellectual disability, neurological symptoms, and behavioral problems.
• The treatment of purine metabolism disorders often involves a combination of medication and lifestyle changes, such as dietary modifications to reduce purine intake and increase hydration.
• In some cases, enzyme replacement therapy or stem cell transplantation is a treatment option.
• Ongoing research is focused on developing new treatments for purine metabolism disorders and gaining a better understanding of their underlying mechanisms.
• Advances in genetic testing have also allowed for early diagnosis and management of purine metabolism disorders, leading to better outcomes for patients.

Frequently Asked Questions (FAQs)

Final words

In conclusion, purine synthesis is a highly regulated process essential for the proper functioning of many biological processes. The production of purine nucleotides is critical for DNA and RNA synthesis and other metabolic pathways and signaling molecules.

The dysregulation of purine metabolism can lead to various disorders, including gout and Lesch-Nyhan syndrome.

In recent years, significant progress has been made in understanding the mechanisms behind purine biosynthesis and its regulation. It has led to the development of novel therapies for diseases associated with purine metabolism, such as gout.

Ongoing research in this area aims further to elucidate the intricate regulatory mechanisms of purine biosynthesis and develop new therapeutic strategies for treating purine metabolism disorders.

The study of purine metabolism remains an active and exciting field with significant potential for future developments.

Understanding the mechanisms behind purine synthesis is critical for developing new treatments for various disorders and expanding our knowledge of the complex processes that govern our biology.