Xanthosine phosphorylase

Last updated
Properties
NameXanthosine Phosphorylase
Gene NamesxapA pndA b2407 JW2398
SynonymspndA, PNPII
Locationcytoplasm
Subunit Composition[XapA]6
Molecular Weight29.835 kD
Map Position[2,522,067 <- 2,522,900]
ResourceIdentifier
Uniprot IDP45563
Uniprot NameXAPA ECOLI
GenBank Gene IDAP009048
Genebank Protein ID1651571
PDB ID 1YR3, 1YQU ,1YQQ
Ecogene IDEG20250
EcocycEG20250
ColiBaseb2407
Kegg Geneb2407
EchoBASE IDEB4152
CCDBXAPA ECOLI
BacMap16130333

Xanthosine phosphorylase, also known as inosine-guanosine phosphorylase, is a catalytic enzyme encoded by the XapA gene in E. coli . The presence of xanthosine is known to induce the synthesis of xanthosine phosphorylase by the XapA gene. The enzyme's main functions are nucleoside phosphorolysis and the synthesis of nucleotides, making it a member of the purine nucleoside phosphorylase group. This protein can degrade all purine nucleosides (including xanthosine) except adenosine, deoxyadenosine, hypoxanthine arabinoside. These degradation reactions are reversible in vitro, however, phosphorolysis dominates in vivo. Xanthosine phosphorylase is localized in the cytoplasm because these degradation functions take place there. Xanthosine phosphorylase preferentially uses the neutral form of xanthosine over its monoanionic form because it prefers to be in a neutral environment.

Contents

Structure

figure 1 Dimer of trimer 1YR3.png
figure 1
figure 2 GUANINE.png
figure 2
Phosphate ion figure 3 PO4 ion.png
Phosphate ion figure 3
figure 4 Xanthosin phosphorylase.png
figure 4
figure 5 2D loop.png
figure 5
figure 6 3D with ligand.png
figure 6

There are three crystal structures. The three structures have been submitted under RSCB protein bank under the codes 1YQU (trigonal form with guanine and phosphate), 1YQQ (small orthorhombic form with guanine and phosphate) and 1YR3 (large orthorhombic form with xanthine and sulfate). (In this section all picture and drawing are created base on 1YQQ, excepte specially noted) Like other Purine nucleoside phosphorylases, xanthosine phosphorylase is a trimer but sometime migrates to a hexamer. The hexamer structure is a combine of two trimers rather than a combine of three dimers, therefore the hexamer is known as "dimer of trimers". [1] Figure 1 shows the "dimer of trimers". The two trimer parts are upper right part and lower left part.

The ligand chemical components of Xanthosine phosphorylase (1YQQ) are a guanine and a phosphate ion.(figure 2 and figure 3) The ligands are buried in the loop. (figure 6) As the Jmol picture shows, the highlighted ligands are buried in the loop of the xanthosine phosphorylase. There are three loop 3D structure in the enzyme, each loop buries a phosphate ion and a guanine.

In the enzyme's crystal structure (figure 4) there is the presence of superimposible hexamers in the crystal forms imply that the hexamer is the predominant form under crystallization conditions. The structure is best described as a dimer of trimers. The 3D structure is shown below. From the 3D structure, the protein domain is a mixed of alpha helix and beta sheet. The alpha helix and beta sheet creates a loop.

The topology diagram (figure 5) shows the connectivity of the secondary structure which is alternative of alpha helices and beta sheets.

History and discovery

In 1980, Hammer-Jepersen K, Buxton RS, and Hansen TD discovered the presence of a second purine nucleoside phosphorylase in wild strains of E. coli K-12 after it demonstrated growth on xanthosine. They came to the conclusion that xanthosine was the only compound that could induce xanthosine phosphorylase. They did not discover any other enzyme through this process, xanthosine phosphorylase was the only new resulting product.

Synthesis

E.coli

Xanthosine phosphorylase(PNP-ĮĮ) is a kind of purine nucleoside phosphorylases (PNPs) in E. coli and it is located in the cytosol. [2] The enzyme is encoded by a 873bp nucleotide gene sequence in E. coli, [3] which encodes XapA, XapB and XapR According to Seeger's research, the XapA gene can't be expressed without the cooperation of XapR gene. XapA gene and XapR gene are almost transferred at the same time. Therefore, the XapA gene and XapB gene is located very closely. However, over expression of XapR doesn't increase of the XapA gene. The XapA gene's expression only increases maxim eight folds. Therefore other factors are also correspond to the expression of XapA. [4] The absence of xanthosine shut off the promoter of XapA, indicating XapA cannot be synthesized in culture that lack xanthosine. [5]

Human

The gene locus of Xanthosine phosphorylase for humans (Homo sapiens) is on chromosome 14q13.1. [6] When disorders in the coding region occur, they will cause PNP deficiency.

Reaction

Starting material

This enzyme catalyzes the phosphorylation and the enzyme's crystal structure. The "dimer of trimers structure" is helpful for creating the correct orientation of starting materials and converting them into intermediates. Xanthosine phosphorylase will bind to the transition state well, and the active site via the Protein Data Bank page, which can point toward critical residues that the enzyme uses. Sulfate ions in the active site allow for properly imposing stress on this substrate making it more likely to undergo the catalysis and to allow for this mechanism to proceed.

figure 8 XapA catalysis.png
figure 8

catalysis

Experimental data has shown that xanthosine phosphorylase performs purine degradation via a substitution of xanthine for an inorganic phosphate group. This enzyme catalyzes the phosphorylation of xanthosine, inosine and guanosine.(figure 8) Xanthosine phosphorylase is a member of the purine nucleoside phosphorylase group, and its main functions are nucleoside phosphorolysis and synthesis of nucleosides. These actions take place in the cytoplasm so xanthosine phosphorylase is located in the cytoplasm. Xanthosine phosphorylase uses orthophosphate to cleave the N-glycosidic bond of ribonucleosides to yield the formation of the corresponding free purine base and ribose 1-phosphate. [7]

Bifunctional catalysis is the prime pathway that this enzyme (and all other enzymes) has used. Acidic and basic residues that are posted toward the outside of the enzyme. This creates an environment that can correctly impose the phosphorylation mechanism. For example, we can take a look at this PDB structure that shows us all relevant interactions maintained in the enzyme's active site.

Xanthosine phosphorylase has the ability to degrade all purine nucleosides (including xanthosine), but cannot cleave adenosine, deoxyadenosine, or hypoxanthine arabinoside. Although this reaction is reversible in vitro, phosphorolysis is dominant in vivo. Additionally, xanthosine phosphorylase prefers to be in a neutral environment so it preferentially uses the neutral form of xanthosine instead of its monoanionic form. Monoanionic forms would create extreme problems in biological settings and catalysis would not be favorable.

pH

The resonance that Xanthosine is able to undergo. At lower pKa's, xanthosine is in the cationic form and although it is well resonated and fairly stable, it will not be found in this way in the body. In the neutral form, the two ring structures are connected. Xanthosine has two carbonyl oxygens, which means that this can allow for the attack of a nucleophile or stabilizing interactions between the oxygen and a hydrogen. Also, the two nitrogen atoms in the ring could potentially act as nucleophiles, but can also be used for stabilizing interactions that will allow for the proper phosphorylation. (Figure 7)

As a cytoplasmic enzyme, functioning under physiological pH, this enzyme is able to perform purine degradation which can be a result of substitution mechanisms xanthine employs.

figure 7 Xanthosine under different pH.png
figure 7

Applications

E.Coli

Using xanthosine containing culture medium can select the growth of E.coli in the presence of S.enterica colony. Because S.enterica contains enzymes that is very similar to E.coli but S.enterica doesn't encode XapA which is responsible for the catalyze of xanthosine and is found in E.coli. [8]

Human

Xanthosine phosphorylase is a purine nucleoside phosphorylase, it involves in the purine metabolism. When human lacks PNP, toxic metabolites, deoxyguanosine triphosphate will accumulate, especially in lymphocytes. The consequence of the accumulation of toxic metabolites is the disordering function of B-cell and T-cell. [9]

Related Research Articles

<span class="mw-page-title-main">Nucleotide</span> Biological molecules constituting nucleic acids

Nucleotides are organic molecules composed of a nitrogenous base, a pentose sugar and a phosphate. They serve as monomeric units of the nucleic acid polymers – deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), both of which are essential biomolecules within all life-forms on Earth. Nucleotides are obtained in the diet and are also synthesized from common nutrients by the liver.

<span class="mw-page-title-main">Xanthine</span> Chemical compound

Xanthine is a purine base found in most human body tissues and fluids, as well as in other organisms. Several stimulants are derived from xanthine, including caffeine, theophylline, and theobromine.

DnaG is a bacterial DNA primase and is encoded by the dnaG gene. The enzyme DnaG, and any other DNA primase, synthesizes short strands of RNA known as oligonucleotides during DNA replication. These oligonucleotides are known as primers because they act as a starting point for DNA synthesis. DnaG catalyzes the synthesis of oligonucleotides that are 10 to 60 nucleotides long, however most of the oligonucleotides synthesized are 11 nucleotides. These RNA oligonucleotides serve as primers, or starting points, for DNA synthesis by bacterial DNA polymerase III. DnaG is important in bacterial DNA replication because DNA polymerase cannot initiate the synthesis of a DNA strand, but can only add nucleotides to a preexisting strand. DnaG synthesizes a single RNA primer at the origin of replication. This primer serves to prime leading strand DNA synthesis. For the other parental strand, the lagging strand, DnaG synthesizes an RNA primer every few kilobases (kb). These primers serve as substrates for the synthesis of Okazaki fragments.

A salvage pathway is a pathway in which a biological product is produced from intermediates in the degradative pathway of its own or a similar substance. The term often refers to nucleotide salvage in particular, in which nucleotides are synthesized from intermediates in their degradative pathway.

<span class="mw-page-title-main">Phosphorylase</span> Class of enzymes

In biochemistry, phosphorylases are enzymes that catalyze the addition of a phosphate group from an inorganic phosphate (phosphate+hydrogen) to an acceptor.

<span class="mw-page-title-main">Glycogen phosphorylase</span> Class of enzymes

Glycogen phosphorylase is one of the phosphorylase enzymes. Glycogen phosphorylase catalyzes the rate-limiting step in glycogenolysis in animals by releasing glucose-1-phosphate from the terminal alpha-1,4-glycosidic bond. Glycogen phosphorylase is also studied as a model protein regulated by both reversible phosphorylation and allosteric effects.

<span class="mw-page-title-main">Succinyl coenzyme A synthetase</span> Class of enzymes

Succinyl coenzyme A synthetase is an enzyme that catalyzes the reversible reaction of succinyl-CoA to succinate. The enzyme facilitates the coupling of this reaction to the formation of a nucleoside triphosphate molecule from an inorganic phosphate molecule and a nucleoside diphosphate molecule. It plays a key role as one of the catalysts involved in the citric acid cycle, a central pathway in cellular metabolism, and it is located within the mitochondrial matrix of a cell.

<span class="mw-page-title-main">Purine nucleoside phosphorylase</span> Enzyme

Purine nucleoside phosphorylase, PNP, PNPase or inosine phosphorylase is an enzyme that in humans is encoded by the NP gene. It catalyzes the chemical reaction

<span class="mw-page-title-main">Nucleic acid metabolism</span> Process

Nucleic acid metabolism is a collective term that refers to the variety of chemical reactions by which nucleic acids are either synthesized or degraded. Nucleic acids are polymers made up of a variety of monomers called nucleotides. Nucleotide synthesis is an anabolic mechanism generally involving the chemical reaction of phosphate, pentose sugar, and a nitrogenous base. Degradation of nucleic acids is a catabolic reaction and the resulting parts of the nucleotides or nucleobases can be salvaged to recreate new nucleotides. Both synthesis and degradation reactions require multiple enzymes to facilitate the event. Defects or deficiencies in these enzymes can lead to a variety of diseases.

<span class="mw-page-title-main">Purine nucleoside phosphorylase deficiency</span> Medical condition

Purine nucleoside phosphorylase deficiency is a rare autosomal recessive metabolic disorder which results in immunodeficiency.

Purine metabolism refers to the metabolic pathways to synthesize and break down purines that are present in many organisms.

<span class="mw-page-title-main">GMP synthase</span>

Guanosine monophosphate synthetase, also known as GMPS is an enzyme that converts xanthosine monophosphate to guanosine monophosphate.

<span class="mw-page-title-main">Polynucleotide phosphorylase</span> Class of enzymes

Polynucleotide Phosphorylase (PNPase) is a bifunctional enzyme with a phosphorolytic 3' to 5' exoribonuclease activity and a 3'-terminal oligonucleotide polymerase activity. That is, it dismantles the RNA chain starting at the 3' end and working toward the 5' end. It also synthesizes long, highly heteropolymeric tails in vivo. It accounts for all of the observed residual polyadenylation in strains of Escherichia coli missing the normal polyadenylation enzyme. Discovered by Marianne Grunberg-Manago working in Severo Ochoa's lab in 1955, the RNA-polymerization activity of PNPase was initially believed to be responsible for DNA-dependent synthesis of messenger RNA, a notion that was disproven by the late 1950s.

<span class="mw-page-title-main">Deoxycytidine kinase</span> Protein-coding gene in the species Homo sapiens

Deoxycytidine kinase (dCK) is an enzyme which is encoded by the DCK gene in humans. dCK predominantly phosphorylates deoxycytidine (dC) and converts dC into deoxycytidine monophosphate. dCK catalyzes one of the initial steps in the nucleoside salvage pathway and has the potential to phosphorylate other preformed nucleosides, specifically deoxyadenosine (dA) and deoxyguanosine (dG), and convert them into their monophosphate forms. There has been recent biomedical research interest in investigating dCK's potential as a therapeutic target for different types of cancer.

In enzymology, a guanosine phosphorylase is an enzyme that catalyzes the chemical reaction

<span class="mw-page-title-main">Thymidine phosphorylase</span> Enzyme

Thymidine phosphorylase is an enzyme that is encoded by the TYMP gene and catalyzes the reaction:

In enzymology, a xanthine phosphoribosyltransferase is an enzyme that catalyzes the chemical reaction

N,N'-diacetylchitobiose phosphorylase is an enzyme with the systematic name N,N'-diacetylchitobiose:phosphate N-acetyl-D-glucosaminyltransferase. This enzyme was found in the genus Vibrio initially but has now been found to be taken up by Escherichia coli as well as many other bacteria. One study shows that Escherichia coli can replicate on a medium that is just composed of GlcNAc a product of phosphorylation of N,N'-diacetylchitobiose as the sole source of carbon. Because E. coli can go on this medium, the enzyme is present. The enzyme has also been found in multiple eukaryotic cells as well, especially in eukaryotes that make chitin and break chitin down. It is believed that N,N'-diacetylchitobiose phosphorylase is an integral part of the phosphoenolpyruvate:glucose phosphotransferase system (PTS). It is assumed that it is involved with Enzyme Complex II of the PTS and is involved with the synthesis of chitin. The enzyme is specific for N,N'-diacetylchitobiose.

Glycogen phosphorylase, liver form (PYGL), also known as human liver glycogen phosphorylase (HLGP), is an enzyme that in humans is encoded by the PYGL gene on chromosome 14. This gene encodes a homodimeric protein that catalyses the cleavage of alpha-1,4-glucosidic bonds to release glucose-1-phosphate from liver glycogen stores. This protein switches from inactive phosphorylase B to active phosphorylase A by phosphorylation of serine residue 14. Activity of this enzyme is further regulated by multiple allosteric effectors and hormonal controls. Humans have three glycogen phosphorylase genes that encode distinct isozymes that are primarily expressed in liver, brain and muscle, respectively. The liver isozyme serves the glycemic demands of the body in general while the brain and muscle isozymes supply just those tissues. In glycogen storage disease type VI, also known as Hers disease, mutations in liver glycogen phosphorylase inhibit the conversion of glycogen to glucose and results in moderate hypoglycemia, mild ketosis, growth retardation and hepatomegaly. Alternative splicing results in multiple transcript variants encoding different isoforms [provided by RefSeq, Feb 2011].

Maltodextrin phosphorylase is a phosphorylase enzyme, more specifically one type of glycosyltransferase. Maltodextrin phosphorylase plays a critical role in maltodextrin metabolism in E. coli. This bacterial enzyme, often referred to as MalP, catalyzes the phosphorolysis of an α-1,4-glycosidic bond in maltodextrins, removing the non-reducing glucosyl residues of linear oligosaccharides as glucose-1-phosphate (Glc1P). Phosphorylases are well-regarded for their allosteric effects on metabolism, however MalP exhibits no allosteric properties. It has a higher affinity for linear oligosaccharides than the related glycogen phosphorylase.

References

  1. ↑ Dandanell, G., Szczepanowski, R. H., Kierdaszuk, B., Shugar, D., & Bochtler, M. (2005). Escherichia coli Purine Nucleoside Phosphorylase II, the Product of the xapA Gene. Journal Of Molecular Biology, 348(1), 113-125. doi:10.1016/j.jmb.2005.02.019
  2. ↑ "Create Account".
  3. ↑ E. coli [ permanent dead link ]
  4. ↑ Seeger, C., Poulsen, C., & Dandanell, G. (1995). Identification and characterization of genes (xapA, xapB, and xapR) involved in xanthosine catabolism in escherichia coli. Journal of Bacteriology, 177(19), 5506-5516.
  5. ↑ Seeger, C., Poulsen, C., & Dandanell, G. (1995). Identification and characterization of genes (xapA, xapB, and xapR) involved in xanthosine catabolism in escherichia coli. Journal of Bacteriology, 177(19), 5506-5516.
  6. ↑ "PNP purine nucleoside phosphorylase [Homo sapiens (Human)] - Gene - NCBI".
  7. ↑ Erion, M. D., & Stoeckler, J. D. (1997). Purine nucleoside phosphorylase. 2. Catalytic mechanism. Biochemistry, 36(39), 11735.
  8. ↑ Hansen, M., Jørgensen, J., & Dandanell, G. (2006). Xanthosine Utilization in Salmonella enterica Serovar Typhimurium Is Recovered by a Single Aspartate-to-Glycine Substitution in Xanthosine Phosphorylase. Journal Of Bacteriology, 188(11), 5. doi:10.1128/JB.01926-05
  9. ↑ Raz Somech, Atar Lev, Amos J. Simon, Suhair Hanna, Amos Etzioni;May 2012;T- and B-cell defects in a novel purine nucleoside phosphorylase mutation; J ALLERGY CLIN IMMUNOL 130:539-542
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