85-32-5

Usage

Uses

Used in Biochemical Research:
5'-Guanylic acid is used as a reagent for chemoand site-specific reductive amination of N2-guanine oligonucleotides, which is essential for various biochemical research applications.
Used as a Flavor Potentiator (Disodium Salt):
The disodium salt of 5'-guanylic acid is used as a flavor potentiator in the food industry. It enhances the umami taste, which is the savory taste found in foods like meat, fish, and vegetables.
Used in the Synthesis of Guanosine Polyphosphates:
5'-Guanylic acid can be further phosphorylated to form guanosine diphosphate (GDP) and guanosine triphosphate (GTP), which are essential for various cellular processes, including protein synthesis, energy transfer, and signal transduction.
Used in the Development of Nucleic Acid Analogues:
5'-Guanylic acid can be used as a building block for the development of nucleic acid analogues, which have potential applications in the treatment of viral infections and genetic disorders.
Used in the Study of Guanylate Cyclase Enzymes:
5'-Guanylic acid is used in the study of guanylate cyclase enzymes, which catalyze the conversion of GTP to cyclic guanosine monophosphate (cGMP). This enzyme plays a crucial role in various physiological processes, including smooth muscle relaxation, ion transport, and cell signaling.

Purification Methods

Crystallise it from water and dry it at 110o. [Beilstein 26 III/IV 3910.]

Upstream product

• 110-86-1 pyridine 
• 770-12-7 O-phenyl phosphorodichloridate 
• 118-00-3 G 
• 362-76-5 2',3'-isopropylideneguanosine 
• 2212-88-6 2-Cyanoethyl phosphate 
• 2304-09-8 2',3'-isopropylidene 5'-GMP 
• 146-91-8 guanosine-5'-diphosphate 
• 102048-77-1 O^2',O^3'-isopropylidene-[5']guanylic acid bis-(4-nitro-phenyl ester) 
• 69281-33-0 Guanosine 5'-monophosphate imidazolide 
• 78681-87-5 Phosphoric acid (3aR,4R,6R,6aR)-6-(2-amino-6-oxo-1,6-dihydro-purin-9-yl)-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethyl ester bis-(2,2,2-tribromo-ethyl) ester 
• 78571-62-7 Phosphoric acid mono-[(2R,3R,4R,5R)-5-(2-amino-6-oxo-1,6-dihydro-purin-9-yl)-3,4-bis-(tert-butyl-dimethyl-silanyloxy)-tetrahydro-furan-2-ylmethyl] ester 
• 7665-99-8 cyclic guanosine 3',5'-monophosphate 
• 4130-19-2 Diguanosine 5'-tetraphosphate 
• 6674-45-9 α,γ-diguanosine 5'-triphosphate 

Downstream Products

• 120209-61-2 [5']guanylic acid mono-cyclohexylamide 
• 146-91-8 guanosine-5'-diphosphate 
• 523-98-8 XMP 
• 86-01-1 Guanosine 5'-triphosphate 
• 89999-10-0 guanosine 5'-monophosphate di-tetramethylammonium salt 
• 79953-11-0 C₁₆H₁₇N₈O₁₀P 
• 79953-10-9 C₁₆H₁₈N₇O₁₁PS 
• 36051-31-7 Guanosine 5'-triphosphate 
• 118-00-3 G 
• 6674-45-9 α,γ-diguanosine 5'-triphosphate 
• 34692-44-9 α,β-diguanosine 5'-diphosphate 
• 80242-42-8 guanosine-5'-(2-methylimidazol-1-yl phosphate) 
• 58-64-0 adenosine 5'-diphosphate 
• 26753-01-5 O¹,O⁵-Diphosphono-α-D-ribofuranose 

Relevant academic research and scientific papers

Biochemical characterization of recombinant guaA-encoded guanosine monophosphate synthetase (EC 6.3.5.2) from Mycobacterium tuberculosis H37Rv strain

Administration of the current tuberculosis (TB) vaccine to newborns is not a reliable route for preventing TB in adults. The conversion of XMP to GMP is catalyzed by guaA-encoded GMP synthetase (GMPS), and deletions in the Shiguella flexneri guaBA operon led to an attenuated auxotrophic strain. Here we present the cloning, expression, and purification of recombinant guaA-encoded GMPS from Mycobacterium tuberculosis (MtGMPS). Mass spectrometry data, oligomeric state determination, steady-state kinetics, isothermal titration calorimetry (ITC), and multiple sequence alignment are also presented. The homodimeric MtGMPS catalyzes the conversion of XMP, MgATP, and glutamine into GMP, ADP, PP i, and glutamate. XMP, NH4+, and Mg2+ displayed positive homotropic cooperativity, whereas ATP and glutamine displayed hyperbolic saturation curves. The activity of ATP pyrophosphatase domain is independent of glutamine amidotransferase domain, whereas the latter cannot catalyze hydrolysis of glutamine to NH3 and glutamate in the absence of substrates. ITC data suggest random order of binding of substrates, and PPi is the last product released. Sequence comparison analysis showed conservation of both Cys-His-Glu catalytic triad of N-terminal Class I amidotransferase and of amino acid residues of the P-loop of the N-type ATP pyrophosphatase family.

Histidine and thermal copolymers of amino acids containing histidine as prebiotic inhibitor for the template-directed formation of oligoguanylate on a poly(C) template

Possibility of the cooperative chemical evolution of nucleic acids and proteins has been investigated using the template-directed formation of oligoguanylate with amino acids and thermal copolymers of amino acids, in which strong inhibition by histidine containing thermal copolymer and histidine itself was observed. The inhibition is regarded as prebiotic enzymatic activities for the hydrolysis of activated nucleotide monomer and the formation of pyrophospho-capped oligoguanylate.

Characterization of complexes of nucleoside-5′-phosphorothioate analogues with zinc ions

On the basis of the high affinity of Zn2+ to sulfur and imidazole, we targeted nucleotides such as GDP-β-S, ADP-β-S, and AP3(β-S)A, as potential biocompatible Zn2+-chelators. The thiophosphate moiety enhanced the stability of the Zn2+- nucleotide complex by about 0.7 log units. ATP-α,β-CH 2-γ-S formed the most stable Zn2+-complex studied here, log K 6.50, being ～0.8 and ～1.1 log units more stable than ATP-γ-S-Zn2+ and ATP-Zn2+ complexes, and was the major species, 84%, under physiological pH. Guanine nucleotides Zn2+ complexes were more stable by 0.3-0.4 log units than the corresponding adenine nucleotide complexes. Likewise, AP3(β-S)A-zinc complex was ～0.5 log units more stable than AP3A complex. 1H- and 31P NMR monitored Zn2+ titration showed that Zn 2+ coordinates with the purine nucleotide N7-nitrogen atom, the terminal phosphate, and the adjacent phosphate. In conclusion, replacement of a terminal phosphate by a thiophosphate group resulted in decrease of the acidity of the phosphate moiety by approximately one log unit, and increase of stability of Zn2+-complexes of the latter analogues by up to 0.7 log units. A terminal phosphorothioate contributed more to the stability of nucleotide-Zn2+ complexes than a bridging phosphorothioate.

High Level Expression of XMP Aminase in Escherichia coli and Its Application for the Industrial Production of 5′-Guanylic Acid

To improve the efficiency of the enzymatic conversion of 5′-xanthylic acid (XMP) to 5′-guanylic acid (GMP), we attempted to increase the activity of the conversion enzyme, XMP aminase (GMP synthetase) encoded by the guaA gene in Escherichia coli. By connecting the PL promoter of λ phage, the SD sequence of trpL of E. coli, and ATG, at a suitable position upstream of the guaA gene, we obtained plasmid pPLA66. Sequencing of the nucleotides of the upstream region of the guaA gene on pPLA66 showed that the C-terminal region of the guaB gene, which encodes IMP dehydrogenase, was conserved and a short peptide consisted of 14 amino acids was coded. E. coli MP347/pPLA66 showed an increase in the activity of approximately 370 times when compared with that of the strain MM294, and the amount of the enzyme protein represented approx. 34% of the total cellular protein. Strain MP347/pPLA66 was cultivated in a 5-liter jar fermentor using a medium which contained mainly corn steep liquor. The culture broth had high XMP aminase activity. In the conversion reaction using mixed broths consisted of 600ml of XMP-fermentation broth of Corynebacterium ammoniagenes KY13203 and 30 ml of cultured broth of E. coli MP347/pPLA66, a surfactant, Nymeen S-215 and xylene were added to the reaction mixture to make the cell membrane permeable to nucleotides. After 23 h of the reaction, 70mg/ml (131 mM) of GMP·Na2·7H2O was accumulated from 83 mg/ml (155 mM) of XMP·Na3 ·7H2O, without addition of ATP. The molar conversion yield was approx. 85%. The facts that the cell membrane was treated to allow nucleotides to permeate and that the conversion reaction proceeded well enough in spite of a small amount of E. coli cells indicate ATP was regenerated from AMP by C. ammoniagenes cells and supplied to E. coli cells. Therefore, it was considered that the coupling reaction between these two kind of strains was established.

Catalytic activity of human guanylate-binding protein 1 coupled to the release of structural restraints imposed by the C-terminal domain

Human guanylate-binding protein 1 (hGBP-1) shows a dimer-induced acceleration of the GTPase activity yielding GDP as well as GMP. While the head-to-head dimerization of the large GTPase (LG) domain is well understood, the role of the rest of the protein, particularly of the GTPase effector domain (GED), in dimerization and GTP hydrolysis is still obscure. In this study, with truncations and point mutations on hGBP-1 and by means of biochemical and biophysical methods, we demonstrate that the intramolecular communication between the LG domain and the GED (LG:GED) is crucial for protein dimerization and dimer-stimulated GTP hydrolysis. In the course of GTP binding and γ-phosphate cleavage, conformational changes within hGBP-1 are controlled by a chain of amino acids ranging from the region near the nucleotide-binding pocket to the distant LG:GED interface and lead to the release of the GED from the LG domain. This opening of the structure allows the protein to form GED:GED contacts within the dimer, in addition to the established LG:LG interface. After releasing the cleaved γ-phosphate, the dimer either dissociates yielding GDP as the final product or it stays dimeric to further cleave the β-phosphate yielding GMP. The second phosphate cleavage step, that is, the formation of GMP, is even more strongly coupled to structural changes and thus more sensitive to structural restraints imposed by the GED. Altogether, we depict a comprehensive mechanism of GTP hydrolysis catalyzed by hGBP-1, which provides a detailed molecular understanding of the enzymatic activity connected to large structural rearrangements of the protein. Database: Structural data are available in RCSB Protein Data Bank under the accession numbers: 1F5N, 1DG3, 2B92.

Helices on Interdomain Interface Couple Catalysis in the ATPPase Domain with Allostery in Plasmodium falciparum GMP Synthetase

GMP synthetase catalyses the conversion of XMP to GMP through a series of reactions that include hydrolysis of Gln to generate ammonia in the glutamine amidotransferase (GATase) domain, activation of XMP to adenyl-XMP intermediate in the ATP pyrophosphatase (ATPPase) domain and reaction of ammonia with the intermediate to generate GMP. The functioning of GMP synthetases entails bidirectional domain crosstalk, which leads to allosteric activation of the GATase domain, synchronization of catalytic events and tunnelling of ammonia. Herein, we have taken recourse to the analysis of structures of GMP synthetases, site-directed mutagenesis and steady-state and transient kinetics on the Plasmodium falciparum enzyme to decipher the molecular basis of catalysis in the ATPPase domain and domain crosstalk. Our results suggest an arrangement at the interdomain interface, of helices with residues that play roles in ATPPase catalysis as well as domain crosstalk enabling the coupling of ATPPase catalysis with GATase activation. Overall, the study enhances our understanding of GMP synthetases, which are drug targets in many infectious pathogens.

Synthesis of ribonucleotides from the corresponding ribonucleosides under plausible prebiotic conditions within self-assembled supramolecular structures

Abiotic synthesis of ribonucleotides, mainly at the 5′ position, from the corresponding ribonucleosides within self-assembled supramolecular structures, based on guanosine:borate hydrogels, was carried out in the temperature range of 70-90 °C, using urea and a phosphate source (K2HPO4 or hydroxyapatite). Phosphorylation is possible at initial concentrations of guanosine lower than 20 mM and it is more efficient using wet/dry cycles. Monoamidophosphate (and, eventually, diamidophosphate), diamidodiphosphate and pyrophosphate are intermediates in the synthesis of ribonucleotides. These conclusions are supported by NMR spectroscopy and mass spectrometry analysis of samples. On the other hand, after reaction, hydrogels can produce globular aggregates by the addition of water and decreasing temperature, thus confirming that ribonucleotides, once activated under suitable conditions, could form polyribonucleotides.

Cloning, expression and biochemical characterization of xanthine and adenine phosphoribosyltransferases from Thermus thermophilus HB8

Purine phosphoribosyltransferases, purine PRTs, are essential enzymes in the purine salvage pathway of living organisms. They are involved in the formation of C-N glycosidic bonds in purine nucleosides-5′-monophosphate (NMPs) through the transfer of the 5-phosphoribosyl group from 5-phospho-α-D-ribosyl-1-pyrophosphate (PRPP) to purine nucleobases in the presence of Mg2+. Herein, we report a simple and thermostable process for the one-pot, one-step synthesis of some purine NMPs using xanthine phosphoribosyltransferase, XPRT or adenine phosphoribosyltransferase, APRT2, from Thermus thermophilus HB8. In this sense, the cloning, expression and purification of TtXPRT and TtAPRT2 is described for the first time. Both genes, xprt and aprt2 were expressed as his-tagged enzymes in E. coli BL21(DE3) and purified by a heat-shock treatment, followed by Ni-affinity chromatography and a final, polishing gel-filtration chromatography. Biochemical characterization revealed TtXPRT as a tetramer and TtAPRT2 as a dimer. In addition, both enzymes displayed a strong temperature dependence (relative activity >75% in a temperature range from 70 to 90 °C), but they also showed very different behaviour under the influence of pH. While TtXPRT is active in a pH range from 5 to 7, TtAPRT2 has a high dependence of alkaline conditions, showing highest activity values in a pH range from 8 to 10. Finally, substrate specificity studies were performed in order to explore their potential as industrial biocatalyst for NMPs synthesis.

Thermophilic phosphoribosyltransferases Thermus thermophilus HB27 in nucleotide synthesis

Phosphoribosyltransferases are the tools that allow the synthesis of nucleotide analogues using multi-enzymatic cascades. The recombinant adenine phosphoribosyltransferase (TthAPRT) and hypoxanthine phosphoribosyltransferase (TthHPRT) from Thermus thermophilus HB27 were expressed in E.coli strains and purified by chromatographic methods with yields of 10-13 mg per liter of culture. The activity dependence of TthAPRT and TthHPRT on different factors was investigated along with the substrate specificity towards different heterocyclic bases. The kinetic parameters for TthHPRT with natural substrates were determined. Two nucleotides were synthesized: 9-(β-D-ribofuranosyl)-2-chloroadenine 5'-monophosphate (2-l-AMP) using TthAPRT and 1-(β-Dribofuranosyl)pyrazolo[3,4-d]pyrimidine-4-one 5'-monophosphate (Allop-MP) using TthPRT.

A unique choanoflagellate enzyme rhodopsin exhibits lightdependent cyclic nucleotide phosphodiesterase activity

Photoactivated adenylyl cyclase (PAC) and guanylyl cyclase rhodopsin increase the concentrations of intracellular cyclic nucleotides upon illumination, serving as promising secondgeneration tools in optogenetics. To broaden the arsenal of such tools, it is desirable to have light-activatable enzymes that can decrease cyclic nucleotide concentrations in cells. Here, we report on an unusual microbial rhodopsin that may be able to meet the demand. It is found in the choanoflagellate Salpingoeca rosetta and contains a C-terminal cyclic nucleotide phosphodiesterase (PDE) domain. We examined the enzymatic activity of the protein (named Rh-PDE) both in HEK293 membranes and whole cells. Although Rh-PDE was constitutively active in the dark, illumination increased its hydrolytic activity 1.4-fold toward cGMP and 1.6-fold toward cAMP, as measured in isolated crude membranes. Purified full-length Rh-PDE displayed maximal light absorption at 492 nm and formed the M intermediate with the deprotonated Schiff base upon illumination. The M state decayed to the parent spectral state in 7 s, producing long-lasting activation of the enzyme domain with increased activity. We discuss a possible mechanism of the Rh-PDE activation by light. Furthermore, Rh-PDE decreased cAMP concentration in HEK293 cells in a light-dependent manner and could do so repeatedly without losing activity. Thus, Rh-PDE may hold promise as a potential optogenetic tool for light control of intracellular cyclic nucleotides (e.g. to study cyclic nucleotide-associated signal transduction cascades).