THERMODYNAMICS OF ADENINE NUCLEOTIDES BINDING TO DUCK NA,K-ATPASE SUGGESTS THE ROLES OF β-PHOSPHATE IN COMPLEX FORMATION AND γ-PHOSPHATE IN CONFORMATIONAL CHANGES

Petrushanko I.Yu.; Mitkevich V.A.; Anashkina A.A.; Klimanova E.A.; Dergousova E.A.; Lopina O.D.; Makarov A.A.

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Журнал

Бюллетень сибирской медицины

2013

Scopus

ВАК

ESCI

Область наук

Биологические науки

Аннотация научной статьи по биологическим наукам, автор научной работы — Petrushanko I. Yu, Mitkevich V. A., Anashkina A. A., Klimanova E. A., Dergousova E. A.

In all animal cells, active transport of sodium and potassium ions across the plasma membrane is facilitated by Na,K-ATPase affecting the cell volume regulation. The transport is accomplished by enzyme conformational changes between two main states, E1 and E2. Na,K-ATPase in the E1 conformation has a high affinity to ATP and Na +, while enzyme in the E2 conformation has a low affinity to ATP and high affinity to K +. It is not clear if binding of ATP to E1 conformation induces some structural changes and what is a role of phosphate groups of ATP in this process. We have used isothermal titration calorimetry to estimate the equilibrium constants, along with the enthalpic and entropic components for AMP, ADP and ATP binding to Na,K-ATPase from duck salt glands at different temperatures. We have found that β-phosphate of ADP involved in complex formation by increasing the affinity to Na,K-ATPase one order of magnitude, while the γ-phosphate of ATP has not effect on the affinity of the enzyme. Furthermore, we demonstrated that ATP, but not ADP, binding to Na,K-ATPase in E1 state generates a conformational change of the enzyme, which is compatible with the movement of 2350 ± 450 Å 2 of molecular surface area from a solvent exposed to a solvent-protected state. We propose that this structural change is caused by the locking of the P domain of Na,K-ATPase to the γ-phosphate of ATP, thereby reducing the exposed surface area of the protein.

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Текст научной работы на тему «THERMODYNAMICS OF ADENINE NUCLEOTIDES BINDING TO DUCK NA,K-ATPASE SUGGESTS THE ROLES OF β-PHOSPHATE IN COMPLEX FORMATION AND γ-PHOSPHATE IN CONFORMATIONAL CHANGES»

Abstracts

THERMODYNAMICS OF ADENINE NUCLEOTIDES BINDING TO DUCK NA,K-ATPASE SUGGESTS THE ROLES OF ß-PHOSPHATE IN COMPLEX FORMATION AND y-PHOSPHATE IN CONFORMATIONAL CHANGES

Petrushanko, I.Yu.1, Mitkevich, V.A.1, Anashkina, A.A.1, Klimanova, E.A2, Dergousova, E.A.2, Lopina, O.D.2, and Makarov, A.A.1

1 Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russian Federation

2 Department of Biochemistry, School of Biology, M. V. Lomonosov Moscow State University, Moscow, Russian Federation

In all animal cells, active transport of sodium and potassium ions across the plasma membrane is facilitated by Na,K-ATPase affecting the cell volume regulation. The transport is accomplished by enzyme conformational changes between two main states, E1 and E2. Na,K-ATPase in the E1 conformation has a high affinity to ATP and Na+, while enzyme in the E2 conformation has a low affinity to ATP and high affinity to K+. It is not clear if binding of ATP to E1 conformation induces some structural changes and what is a role of phosphate groups of ATP in this process. We have used isothermal titration calorimetry to estimate the equilibrium constants, along with the enthalpic and entropic components for AMP, ADP and ATP binding to Na,K-ATPase from duck salt glands at different temperatures. We have found that p-

phosphate of ADP involved in complex formation by increasing the affinity to Na,K-ATPase one order of magnitude, while the y-phosphate of ATP has not effect on the affinity of the enzyme. Furthermore, we demonstrated that ATP, but not ADP, binding to Na,K-ATPase in E1 state generates a conformational change of the enzyme, which is compatible with the movement of 2350 ± 450 Â2 of molecular surface area from a solvent exposed to a solvent-protected state. We propose that this structural change is caused by the locking of the P domain of Na,K-ATPase to the y-phosphate of ATP, thereby reducing the exposed surface area of the protein.

Study was supported by RFBR grant №12-04-00103a and State Agreement № 16.512.11.2280.

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Бюллетень сибирской медицины, 2013, том 12, № 4, с. 24-68

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THERMODYNAMICS OF ADENINE NUCLEOTIDES BINDING TO DUCK NA,K-ATPASE SUGGESTS THE ROLES OF β-PHOSPHATE IN COMPLEX FORMATION AND γ-PHOSPHATE IN CONFORMATIONAL CHANGES Текст научной статьи по специальности «Биологические науки»

Аннотация научной статьи по биологическим наукам, автор научной работы — Petrushanko I. Yu, Mitkevich V. A., Anashkina A. A., Klimanova E. A., Dergousova E. A.

Похожие темы научных работ по биологическим наукам , автор научной работы — Petrushanko I. Yu, Mitkevich V. A., Anashkina A. A., Klimanova E. A., Dergousova E. A.

Текст научной работы на тему «THERMODYNAMICS OF ADENINE NUCLEOTIDES BINDING TO DUCK NA,K-ATPASE SUGGESTS THE ROLES OF β-PHOSPHATE IN COMPLEX FORMATION AND γ-PHOSPHATE IN CONFORMATIONAL CHANGES»