| Autor: |
Stachelska-Wierzchowska A; Department of Physics and Biophysics, Faculty of Food Sciences, University of Warmia and Mazury in Olsztyn, 4 Oczapowskiego St., PL-10-719 Olsztyn, Poland., Narczyk M; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, ul. Pasteura 5, PL-02-093 Warsaw, Poland., Wierzchowski J; Department of Physics and Biophysics, Faculty of Food Sciences, University of Warmia and Mazury in Olsztyn, 4 Oczapowskiego St., PL-10-719 Olsztyn, Poland., Bzowska A; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, ul. Pasteura 5, PL-02-093 Warsaw, Poland., Wielgus-Kutrowska B; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, ul. Pasteura 5, PL-02-093 Warsaw, Poland. |
| Jazyk: |
angličtina |
| Zdroj: |
International journal of molecular sciences [Int J Mol Sci] 2024 Sep 27; Vol. 25 (19). Date of Electronic Publication: 2024 Sep 27. |
| DOI: |
10.3390/ijms251910426 |
| Abstrakt: |
Fluorescent markers play important roles in spectroscopic and microscopic research techniques and are broadly used in basic and applied sciences. We have obtained markers with fluorescent properties, two etheno derivatives of 2-aminopurine, as follows: 1,N 2 -etheno-2-aminopurine (1,N 2 -ε2APu, I ) and N 2 ,3-etheno-2-aminopurine (N 2 ,3-ε2APu, II ). In the present paper, we investigate their interaction with two key enzymes of purine metabolism, purine nucleoside phosphorylase (PNP), and xanthine oxidase (XO), using diffraction of X-rays on protein crystals, isothermal titration calorimetry, and fluorescence spectroscopy. Crystals were obtained and structures were solved for WT PNP and D204N-PNP mutant in a complex with N 2 ,3-ε2APu ( II ). In the case of WT PNP-1,N 2 -ε2APu ( I ) complex, the electron density corresponding to the ligand could not be identified in the active site. Small electron density bobbles may indicate that the ligand binds to the active site of a small number of molecules. On the basis of spectroscopic studies in solution, we found that, in contrast to PNP, 1,N 2 -ε2APu ( I ) is the ligand with better affinity to XO. Enzymatic oxidation of ( I ) leads to a marked increase in fluorescence near 400 nm. Hence, we have developed a new method to determine XO activity in biological material, particularly suitable for milk analysis. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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