Strontium Binding to α-Parvalbumin, a Canonical Calcium-Binding Protein of the “EF-Hand” Family
Strontium salts are used for treatment of osteoporosis and bone cancer, but their impact on calcium-mediated physiological processes remains obscure. To explore Sr<sup>2+</sup> interference with Ca<sup>2+</sup> binding to proteins of the EF-hand family, we studied Sr<sup&g...
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2021-08-01
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| Series: | Biomolecules |
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| author | Alisa A. Vologzhannikova Marina P. Shevelyova Alexey S. Kazakov Andrey S. Sokolov Nadezhda I. Borisova Eugene A. Permyakov Nikoleta Kircheva Valya Nikolova Todor Dudev Sergei E. Permyakov |
| author_facet | Alisa A. Vologzhannikova Marina P. Shevelyova Alexey S. Kazakov Andrey S. Sokolov Nadezhda I. Borisova Eugene A. Permyakov Nikoleta Kircheva Valya Nikolova Todor Dudev Sergei E. Permyakov |
| author_sort | Alisa A. Vologzhannikova |
| collection | DOAJ |
| description | Strontium salts are used for treatment of osteoporosis and bone cancer, but their impact on calcium-mediated physiological processes remains obscure. To explore Sr<sup>2+</sup> interference with Ca<sup>2+</sup> binding to proteins of the EF-hand family, we studied Sr<sup>2+</sup>/Ca<sup>2+</sup> interaction with a canonical EF-hand protein, α-parvalbumin (α-PA). Evaluation of the equilibrium metal association constants for the active Ca<sup>2+</sup> binding sites of recombinant human α-PA (‘CD’ and ‘EF’ sites) from fluorimetric titration experiments and isothermal titration calorimetry data gave 4 × 10<sup>9</sup> M<sup>−1</sup> and 4 × 10<sup>9</sup> M<sup>−1</sup> for Ca<sup>2+</sup>, and 2 × 10<sup>7</sup> M<sup>−1</sup> and 2 × 10<sup>6</sup> M<sup>−1</sup> for Sr<sup>2+</sup>. Inactivation of the EF site by homologous substitution of the Ca<sup>2+</sup>-coordinating Glu in position 12 of the EF-loop by Gln decreased Ca<sup>2+</sup>/Sr<sup>2+</sup> affinity of the protein by an order of magnitude, whereas the analogous inactivation of the CD site induced much deeper suppression of the Ca<sup>2+</sup>/Sr<sup>2+</sup> affinity. These results suggest that Sr<sup>2+</sup> and Ca<sup>2+</sup> bind to CD/EF sites of α-PA and the Ca<sup>2+</sup>/Sr<sup>2+</sup> binding are sequential processes with the CD site being occupied first. Spectrofluorimetric Sr<sup>2+</sup> titration of the Ca<sup>2+</sup>-loaded α-PA revealed presence of secondary Sr<sup>2+</sup> binding site(s) with an apparent equilibrium association constant of 4 × 10<sup>5</sup> M<sup>−1</sup>. Fourier-transform infrared spectroscopy data evidence that Ca<sup>2+</sup>/Sr<sup>2+</sup>-loaded forms of α-PA exhibit similar states of their COO<sup>−</sup> groups. Near-UV circular dichroism (CD) data show that Ca<sup>2+</sup>/Sr<sup>2+</sup> binding to α-PA induce similar changes in symmetry of microenvironment of its Phe residues. Far-UV CD experiments reveal that Ca<sup>2+</sup>/Sr<sup>2+</sup> binding are accompanied by nearly identical changes in secondary structure of α-PA. Meanwhile, scanning calorimetry measurements show markedly lower Sr<sup>2+</sup>-induced increase in stability of tertiary structure of α-PA, compared to the Ca<sup>2+</sup>-induced effect. Theoretical modeling using Density Functional Theory computations with Polarizable Continuum Model calculations confirms that Ca<sup>2+</sup>-binding sites of α-PA are well protected against exchange of Ca<sup>2+</sup> for Sr<sup>2+</sup> regardless of coordination number of Sr<sup>2+</sup>, solvent exposure or rigidity of sites. The latter appears to be a key determinant of the Ca<sup>2+</sup>/Sr<sup>2+</sup> selectivity. Overall, despite lowered affinity of α-PA to Sr<sup>2+</sup>, the latter competes with Ca<sup>2+</sup> for the same EF-hands and induces similar structural rearrangements. The presence of a secondary Sr<sup>2+</sup> binding site(s) could be a factor contributing to Sr<sup>2+</sup> impact on the functional activity of proteins. |
| first_indexed | 2024-03-10T08:58:45Z |
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| institution | Directory Open Access Journal |
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| language | English |
| last_indexed | 2024-03-10T08:58:45Z |
| publishDate | 2021-08-01 |
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| series | Biomolecules |
| spelling | doaj.art-bd844b9458e14347a9e48bc9f3e29bbd2023-11-22T06:55:40ZengMDPI AGBiomolecules2218-273X2021-08-01118115810.3390/biom11081158Strontium Binding to α-Parvalbumin, a Canonical Calcium-Binding Protein of the “EF-Hand” FamilyAlisa A. Vologzhannikova0Marina P. Shevelyova1Alexey S. Kazakov2Andrey S. Sokolov3Nadezhda I. Borisova4Eugene A. Permyakov5Nikoleta Kircheva6Valya Nikolova7Todor Dudev8Sergei E. Permyakov9Institute for Biological Instrumentation, Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences, Pushchino, 142290 Moscow, RussiaInstitute for Biological Instrumentation, Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences, Pushchino, 142290 Moscow, RussiaInstitute for Biological Instrumentation, Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences, Pushchino, 142290 Moscow, RussiaInstitute for Biological Instrumentation, Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences, Pushchino, 142290 Moscow, RussiaInstitute for Biological Instrumentation, Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences, Pushchino, 142290 Moscow, RussiaInstitute for Biological Instrumentation, Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences, Pushchino, 142290 Moscow, RussiaInstitute of Optical Materials and Technologies “Acad. J. Malinowski”, Bulgarian Academy of Sciences, 1113 Sofia, BulgariaFaculty of Chemistry and Pharmacy, Sofia University “St. Kl. Ohridski”, 1164 Sofia, BulgariaFaculty of Chemistry and Pharmacy, Sofia University “St. Kl. Ohridski”, 1164 Sofia, BulgariaInstitute for Biological Instrumentation, Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences, Pushchino, 142290 Moscow, RussiaStrontium salts are used for treatment of osteoporosis and bone cancer, but their impact on calcium-mediated physiological processes remains obscure. To explore Sr<sup>2+</sup> interference with Ca<sup>2+</sup> binding to proteins of the EF-hand family, we studied Sr<sup>2+</sup>/Ca<sup>2+</sup> interaction with a canonical EF-hand protein, α-parvalbumin (α-PA). Evaluation of the equilibrium metal association constants for the active Ca<sup>2+</sup> binding sites of recombinant human α-PA (‘CD’ and ‘EF’ sites) from fluorimetric titration experiments and isothermal titration calorimetry data gave 4 × 10<sup>9</sup> M<sup>−1</sup> and 4 × 10<sup>9</sup> M<sup>−1</sup> for Ca<sup>2+</sup>, and 2 × 10<sup>7</sup> M<sup>−1</sup> and 2 × 10<sup>6</sup> M<sup>−1</sup> for Sr<sup>2+</sup>. Inactivation of the EF site by homologous substitution of the Ca<sup>2+</sup>-coordinating Glu in position 12 of the EF-loop by Gln decreased Ca<sup>2+</sup>/Sr<sup>2+</sup> affinity of the protein by an order of magnitude, whereas the analogous inactivation of the CD site induced much deeper suppression of the Ca<sup>2+</sup>/Sr<sup>2+</sup> affinity. These results suggest that Sr<sup>2+</sup> and Ca<sup>2+</sup> bind to CD/EF sites of α-PA and the Ca<sup>2+</sup>/Sr<sup>2+</sup> binding are sequential processes with the CD site being occupied first. Spectrofluorimetric Sr<sup>2+</sup> titration of the Ca<sup>2+</sup>-loaded α-PA revealed presence of secondary Sr<sup>2+</sup> binding site(s) with an apparent equilibrium association constant of 4 × 10<sup>5</sup> M<sup>−1</sup>. Fourier-transform infrared spectroscopy data evidence that Ca<sup>2+</sup>/Sr<sup>2+</sup>-loaded forms of α-PA exhibit similar states of their COO<sup>−</sup> groups. Near-UV circular dichroism (CD) data show that Ca<sup>2+</sup>/Sr<sup>2+</sup> binding to α-PA induce similar changes in symmetry of microenvironment of its Phe residues. Far-UV CD experiments reveal that Ca<sup>2+</sup>/Sr<sup>2+</sup> binding are accompanied by nearly identical changes in secondary structure of α-PA. Meanwhile, scanning calorimetry measurements show markedly lower Sr<sup>2+</sup>-induced increase in stability of tertiary structure of α-PA, compared to the Ca<sup>2+</sup>-induced effect. Theoretical modeling using Density Functional Theory computations with Polarizable Continuum Model calculations confirms that Ca<sup>2+</sup>-binding sites of α-PA are well protected against exchange of Ca<sup>2+</sup> for Sr<sup>2+</sup> regardless of coordination number of Sr<sup>2+</sup>, solvent exposure or rigidity of sites. The latter appears to be a key determinant of the Ca<sup>2+</sup>/Sr<sup>2+</sup> selectivity. Overall, despite lowered affinity of α-PA to Sr<sup>2+</sup>, the latter competes with Ca<sup>2+</sup> for the same EF-hands and induces similar structural rearrangements. The presence of a secondary Sr<sup>2+</sup> binding site(s) could be a factor contributing to Sr<sup>2+</sup> impact on the functional activity of proteins.https://www.mdpi.com/2218-273X/11/8/1158metal selectivitycalcium-binding proteinsEF-handparvalbuminprotein structureprotein stability |
| spellingShingle | Alisa A. Vologzhannikova Marina P. Shevelyova Alexey S. Kazakov Andrey S. Sokolov Nadezhda I. Borisova Eugene A. Permyakov Nikoleta Kircheva Valya Nikolova Todor Dudev Sergei E. Permyakov Strontium Binding to α-Parvalbumin, a Canonical Calcium-Binding Protein of the “EF-Hand” Family Biomolecules metal selectivity calcium-binding proteins EF-hand parvalbumin protein structure protein stability |
| title | Strontium Binding to α-Parvalbumin, a Canonical Calcium-Binding Protein of the “EF-Hand” Family |
| title_full | Strontium Binding to α-Parvalbumin, a Canonical Calcium-Binding Protein of the “EF-Hand” Family |
| title_fullStr | Strontium Binding to α-Parvalbumin, a Canonical Calcium-Binding Protein of the “EF-Hand” Family |
| title_full_unstemmed | Strontium Binding to α-Parvalbumin, a Canonical Calcium-Binding Protein of the “EF-Hand” Family |
| title_short | Strontium Binding to α-Parvalbumin, a Canonical Calcium-Binding Protein of the “EF-Hand” Family |
| title_sort | strontium binding to α parvalbumin a canonical calcium binding protein of the ef hand family |
| topic | metal selectivity calcium-binding proteins EF-hand parvalbumin protein structure protein stability |
| url | https://www.mdpi.com/2218-273X/11/8/1158 |
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