Development of a Simple Assay Method for Adenosine Deaminase via Enzymatic Formation of an Inosine-Tb3+ Complex

Adenosine deaminase (ADA), which catalyzes the irreversible deamination of adenosine to inosine, is related to various human diseases such as tuberculous peritonitis and leukemia. Therefore, the method used to detect ADA activity and screen the effectiveness of various inhibitor candidates has important implications for the diagnosis treatment for various human diseases. A simple and rapid assay method for ADA, based on the enzymatic formation of a luminescent lanthanide complex, is proposed in this study. Inosine, an enzymatic product of ADA with stronger sensitization efficiency for Tb3+ than adenosine, produced a strong luminescence by forming an inosine-Tb3+ complex, and it enabled the direct monitoring of ADA activity in real-time. By introducing only Tb3+ to adenosine and ADA in the buffer, the enhancement of luminescence enabled the detection of a low concentration of ADA (detection limit 1.6 U/L). Moreover, this method could accurately determine the inhibition efficiency (IC50) of the known ADA inhibitor, erhythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), and the inhibition of ADA could be confirmed by the naked eye. Considering its simplicity, this assay could be extended to the high-throughput screening of various ADA inhibitor candidates.

[1]  Elias Pershagen,et al.  Multiplex detection of enzymatic activity with responsive lanthanide-based luminescent probes. , 2015, Angewandte Chemie.

[2]  T. Gunnlaugsson,et al.  Lanthanide luminescent displacement assays: the sensing of phosphate anions using Eu(III)-cyclen-conjugated gold nanoparticles in aqueous solution. , 2008, Journal of the American Chemical Society.

[3]  Gianluca Accorsi,et al.  Design of luminescent lanthanide complexes: From molecules to highly efficient photo-emitting materials , 2010 .

[4]  Massimo Guardigli,et al.  Luminescent lanthanide complexes as photochemical supramolecular devices , 1993 .

[5]  J. Zweier,et al.  Adenosine Deaminase Inhibition Prevents Free Radical-mediated Injury in the Postischemic Heart (*) , 1996, The Journal of Biological Chemistry.

[6]  Li Wang,et al.  A terbium chelate based fluorescent assay for alkaline phosphatase in biological fluid , 2014 .

[7]  H. Harris,et al.  Further data on the adenosine deaminase (ADA) polymorphism and a report of a new phenotype , 1969, Annals of human genetics.

[8]  D. Martin,et al.  Deoxyadenosine triphosphate as a potentially toxic metabolite in adenosine deaminase deficiency. , 1978, Proceedings of the National Academy of Sciences of the United States of America.

[9]  Wei Huang,et al.  Recent developments in lanthanide-based luminescent probes , 2014 .

[10]  Qing-Ying Luo,et al.  Graphene oxide based fluorescent aptasensor for adenosine deaminase detection using adenosine as the substrate. , 2012, Biosensors & bioelectronics.

[11]  R. Hirschhorn.  Adenosine deaminase deficiency: molecular basis and recent developments. , 1995, Clinical immunology and immunopathology.

[12]  I. A. A. El-Gawad,et al.  Comparison of the Coordination Tendency of Amino Acids, Nucleobases, or Mononucleotides Toward the Monomeric and Dimeric Lanthanide Complexes with Biologically Important Compounds , 2011 .

[13]  J. Hutton,et al.  Overproduction of adenine deoxynucleosides and deoxynucletides in adenosine deaminase deficiency with severe combined immunodeficiency disease. , 1978, The Journal of clinical investigation.

[14]  K. Raymond,et al.  From antenna to assay: lessons learned in lanthanide luminescence. , 2009, Accounts of chemical research.

[15]  V. Pierre,et al.  A ratiometric probe for the selective time-gated luminescence detection of potassium in water. , 2011, Chemical communications.

[16]  J. Ungerer,et al.  Kinetic determination of serum adenosine deaminase. , 1993, Clinical chemistry.

[17]  S. Achilefu,et al.  Fluorescence lifetime measurements and biological imaging. , 2010, Chemical reviews.

[18]  M. Pelegrí,et al.  Adenosine deaminase in body fluids: a useful diagnostic tool in tuberculosis. , 1989, Clinical biochemistry.

[19]  R. Stern,et al.  Purine nucleoside metabolism in the erythrocytes of patients with adenosine deaminase deficiency and severe combined immunodeficiency. , 1976, The Journal of clinical investigation.

[20]  A. Saboury,et al.  A Kinetic Comparison on the Inhibition of Adenosine Deaminase by Purine Drugs , 2007 .

[21]  Genxi Li,et al.  Strategy to fabricate an electrochemical aptasensor: application to the assay of adenosine deaminase activity. , 2010, Analytical chemistry.

[22]  E. Slominska,et al.  Adenosine deaminase inhibition suppresses progression of 4T1 murine breast cancer by adenosine receptor‐dependent mechanisms , 2018, Journal of cellular and molecular medicine.

[23]  Yi Lin,et al.  A gold nanoparticle-based label free colorimetric aptasensor for adenosine deaminase detection and inhibition assay. , 2015, The Analyst.

[24]  Kai Zhang,et al.  A new strategy based on aptasensor to time-resolved fluorescence assay for adenosine deaminase activity. , 2013, Biosensors & bioelectronics.

[25]  Nathaniel L Rosi,et al.  Near-infrared luminescent lanthanide MOF barcodes. , 2009, Journal of the American Chemical Society.

[26]  Shulin Zhao,et al.  A carbon nanotubes based fluorescent aptasensor for highly sensitive detection of adenosine deaminase activity and inhibitor screening in natural extracts. , 2014, Journal of pharmaceutical and biomedical analysis.

[27]  J. Smyth,et al.  Correlation of adenosine deaminase activity with cell surface markers in acute lymphoblastic leukemia. , 1978, Journal of Clinical Investigation.

[28]  L. Burgess,et al.  The use of adenosine deaminase as a diagnostic tool for peritoneal tuberculosis. , 2001, Tuberculosis.

[29]  G. Cristalli,et al.  Adenosine deaminase: Functional implications and different classes of inhibitors , 2001, Medicinal research reviews.

[30]  Yanli Tang,et al.  Water-soluble conjugated polymer as a platform for adenosine deaminase sensing based on fluorescence resonance energy transfer technique. , 2014, Analytical chemistry.

[31]  J. Hutton,et al.  Identification and quantitation of adenine deoxynucleotides in erythrocytes of a patient with adenosine deaminase deficiency and severe combined immunodeficiency. , 1978, The Journal of biological chemistry.

[32]  B. Graham,et al.  Lanthanide complexes for luminescence-based sensing of low molecular weight analytes , 2017, Coordination Chemistry Reviews.

[33]  K. Kikuchi,et al.  Lanthanide-based protease activity sensors for time-resolved fluorescence measurements. , 2008, Journal of the American Chemical Society.

[34]  Alessandro Aiuti,et al.  Gene therapy for immunodeficiency due to adenosine deaminase deficiency. , 2009, The New England journal of medicine.

[35]  P. Daddona,et al.  Human Adenosine Deaminase Binding Protein , 2011 .

[36]  J. Georges,et al.  Lanthanide-sensitized luminescence and applications to the determination of organic analytes : a review , 1993 .

[37]  J. Bünzli,et al.  Taking advantage of luminescent lanthanide ions. , 2005, Chemical Society reviews.

[38]  S. Petoud,et al.  Zinc-adeninate metal-organic framework for aqueous encapsulation and sensitization of near-infrared and visible emitting lanthanide cations. , 2011, Journal of the American Chemical Society.

[39]  M. Weyden,et al.  Human adenosine deaminase. Distribution and properties. , 1976, The Journal of biological chemistry.

[40]  G. Janossy,et al.  Enzyme and membrane markers in leukaemia: recent developments. , 1981, Journal of clinical pathology.