dATP: function, structure & key applications

Introduction to dATP

Deoxyadenosine triphosphate (dATP) is one of four nucleotides that play an important role in DNA synthesis, replication and repair. Every time a cell divides, DNA polymerases are required to duplicate the cell’s DNA, so that a copy of the original DNA molecule can be passed to each daughter cell. Therefore DNA polymerase adds nucleotides (dATP, dCTP, dTTP, dGTP) In this way, genetic information is passed down from generation to generation. All of the nucleotides consist of a deoxyribose sugar, an base and three phosphate groups. In case of dATP the base is adenine. As one of the four building blocks of DNA, dATP is widely used in genetic research, diagnostics and biotechnology. E.g. in all molecular biology applications including PCR, real-time PCR, high fidelity and long range PCR, Loop-mediated isothermal amplification (LAMP), cDNA synthesis, RT-PCR, RDA, MDA, DNA labelling and DNA sequencing.

What is dATP?

The structure of dATP

The molecular structure of dATP consists of three major components:

1. Three phosphate groups:

Provide the energy necessary to incorporate dATP into the growing DNA strand during synthesis. DNA polymerase covalently links nucleotide to nucleotide. As the nucleotide chain grows, the free -OH group of the sugar at the 3′ carbon is linked to the α-phosphate at the 5′ carbon of the next (d)NTP, releasing the β- and γ-phosphate groups as pyrophosphate (PPi).

2. Deoxyribose sugar:

The sugar of dATP, deoxyribose, looks like a five-sided (furanose) ring in which four of the five carbon atoms and one oxygen atom are part of the ring. The fifth carbon is a branch of the ring. In nucleic acid synthesis, the 3’OH of a growing chain of e.g. dATP binds to the α-phosphate of another nucleotide through phosphodiester bonds. Both the sugar and the phosphate group form the backbone of DNA and bind to other nucleotides.

3. Adenine base (short A):

A purine base that specifically binds to thymin in DNA or uridine in RNA, thereby ensuring the stability of the double helix. Adenine has a central role in cellular respiration. It is part of adenosine triphosphate (dATP) which provides the energy for protein synthesis, muscle contraction and nerve impulse propagation.

Role of dATP in DNA synthesis and replication

DNA replication process

During DNA replication, dATP serves as a substrate for DNA polymerase in the elongation process.

dATP is incorporated into the new DNA strand as a nucleotide complementary to thymine. This specific binding to thymin in DNA is called complementary base pairing. Thereby, adenine in dATP forms hydrogen bonds with thymine in dTTP, ensuring the accuracy of replication.

DNA repair and integrity

dATP also plays an essential role in DNA repair by preventing mutations. Through proper incorporation during replication, dATP contributes to genetic stability. By supporting repair processes dATP is involved in mechanisms that correct damaged DNA bases.

Importance of dATP in genetic and molecular research

Genetic studies and therapeutics

dATP is gaining attention in the pharmaceutical industry because of its potential therapeutic benefits and its relation in genetics and gene editing. Research suggests that dATP may play a role in improving cellular metabolism and energy production, making it a candidate for the treatment of several conditions, including mitochondrial diseases (read more on the MITGEST project website) and age-related disorders. E.g. it is known that the dATP-dependent and -independent release of cytochrome c from mitochondria cannot be inhibited by the protease inhibitor z-VAD. It has long been investigated whether a chemical compound could directly cause the release of cytochrome c from mitochondria and explain the toxicity of dATP in the genetic diseases such as adenosine deaminase deficiency and purine nucleotide phosphorylase deficiency. Such mutations or alterations in the genomic loci can be identified by genetic markers. It is a gene or short DNA sequence, such as a sequence surrounding a single base-pair change or a long one, like minisatellites.

Applications in research

dATP is one of four DNA precursor and is widespread in laboratories for studies involving genetic stability, gene expression, etc. E.g. the intracellular concentrations of the four deoxyribonucleoside triphosphates dATP, dGTP, dTTP, dCTP are tightly controlled. Imbalances in the four dNTP pools can cause genotoxic consequences. Excess of e.g. dATP can lead to replication errors and mutations by forming incorrect base pairs or forcing DNA chain extension past mismatches before DNA polymerase corrects them via 3′ exonuclease proofreading.

dATP in laboratory settings

Commercial availability and types of dATP products

baseclick offers a variety of modified dATP products developed for specific research applications:

BCT-21 Alkyne-dATP (7-deaza-7-Ethynyl-dATP)

Alkyne-dATP (7-deaza-7-ethynyl-dATP) is easily incorporated into DNA by PCR in place of dATP using polymerases such as Pwo, Deep Vent exo- or KOD XL. The resulting stable and modified PCR fragment is now ready for further functionalisation by click chemistry. We recommend 1-10% dATP substitution.

CAS number: 1208963-36-3 (free acid)

Concentration: 10 mM solution in water

Purity: ≥ 95% (HPLC)

BCT-25 3′-Azido-2′,3′-ddATP

This azide-modified nucleotide closely resembles a natural nucleotide and allows chemoenzymatic labelling of mRNA or ssDNA, e.g. for cDNA synthesis. This dATP variant is also used in our NGS kit such as the ClickSeq Library Prep Kit and the Poly(A)-ClickSeq Library Prep Kit. Due to the small size of the azide group, 3′-Azido-2′,3′-ddATP is well accepted by T7 RNA polymerase, poly(A) polymerase or terminal deoxynucleotidyl transferase (Tdt) for enzymatic incorporation. This is highly advantageous as the labelling of mRNA or ssDNA with our approach is modular and not limited to the enzymatic compatibility of bulky pre-labelled nucleotides.

CAS number: 1383937-03-8 (sodium salt); 92562-94-2 (free acid)

Concentration: 100 mM clear aquaeous solution; colorless

Purity: ≥ 95% (HPLC)

BCT-34 2′-Azido-2′-dATP

2′-Azido-2′-dATP offers modular, chemoenzymatic DNA or RNA labeling, allowing flexible alkyne or DBCO-based bioconjugation at the sugar, ideal for creating labeled DNA or RNA without complex protocols.

CAS number: 69093-69-2

Concentration: colorless solution (100 mM, pH 7.5)

Purity: ≥ 95% (HPLC)

Storage and handling guidelines

dATP is used in all molecular biology applications including PCR, real-time PCR, high fidelity and long range PCR, LAMP, cDNA synthesis, RT-PCR, RDA, MDA, DNA labelling and DNA sequencing. dATP (2′-deoxyadenosine-5′-triphosphate) is an extremely stable nucleotide and usually supplied as a 100 mM aqueous solution with a pH of 7.3 to 7.5. For laboratory use the purity should be over 99% and free from nuclease activity, human DNA and E. coli DNA. dATP is shipped on gel packs to ensure stability during transit. For optimal storage conditions, dATP should be stored at -20 °C. Short-term exposure to ambient temperature, up to one week cumulatively, is possible without compromising the dATP’s integrity. The shelf life of dATP is normally 12 months from the date of delivery. The molecular formula of dATP is C₁₀H₁₆N₅O₁₂P₃ (free acid), with a molecular weight of 491.18g/mol (free acid). The spectroscopic properties of dATP include an absorption maximum (λabs) at 259nm, with an extinction coefficient (ε) of 15.1 L mmol-1 cm-1 in Tris-HCl buffer at pH 7.0.

Future of dATP in biotechnology

Advancements in research and drug development

dNTPs (dATP, dCTP, dTTP, dGTP) are key molecules in all molecular biology applications leading to new scientific discoveries and therapeutic treatments. These include PCR, real-time PCR, high fidelity and long-range PCR, LAMP, cDNA synthesis, RT-PCR, RDA, MDA, DNA labelling and DNA sequencing. One of the key advantages of using dATP in drug delivery systems is its ability to be incorporated into DNA and RNA molecules, allowing targeted delivery of genetic material. This can be particularly useful in gene therapy, where the aim is to introduce or correct specific genes into a patient’s cells. By using dATP-modified nucleotides, such as those offered by baseclick, researchers can create more stable and efficient delivery vehicles that can navigate the complex cellular environment and reach their intended targets.