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Role of potassium levels in pkBADH heterogeneity of NAD+ binding site

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Abstract

Betaine aldehyde dehydrogenase (BADH) catalyzes the oxidation of betaine aldehyde to glycine betaine using NAD+ as a coenzyme. Studies in porcine kidney BADH (pkBADH) suggested that the enzyme exhibits heterogeneity of active sites and undergoes potassium-induced conformational changes. This study aimed to analyze if potassium concentration plays a role in the heterogeneity of pkBADH active sites through changes in NAD+ affinity constants, in its secondary structure content and stability. The enzyme was titrated with NAD+ 1 mM at fixed-variable KCl concentration, and the interaction measured by Isothermal Titration Calorimetry (ITC) and Circular Dichroism (CD). ITC data showed that K+ increased the first active site affinity in a manner dependent on its concentration; KD values to the first site were 14.4, 13.1, and 10.4 μM, at 25, 50, and 75 mM KCl. ΔG values showed that the coenzyme binding is a spontaneous reaction without changes between active sites or depending on KCl concentration. ΔH and TΔSb values showed that NAD+ binding to the active site is an endothermic process and is carried out at the expense of changes in entropy. α-Helix content increased as KCl increased, enzyme (Tm)app values were 2.6 °C and 3.3 °C higher at 20 mM and 200 mM K+. PkBADH molecular model showed three different interaction K+ sites. Results suggested K+ can interact with pkBADH and cause changes in the secondary structure, it provokes changes in the enzyme affinity by the coenzyme, and in the thermostability.

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Acknowledgments

César Muñoz-Bacasehua gratefully acknowledges a scholarship from CONACyT for graduate studies. Authors are grateful to Dr. G. Garza-Ramos for providing facilities and constant support to carry out CD spectra.

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Authors and Affiliations

  1. Centro de Investigación en Alimentación y Desarrollo A.C, GE Astiazaran Rosas 46, 83304, Hermosillo, Sonora, Mexico

    César Muñoz-Bacasehua & Elisa M. Valenzuela-Soto

  2. Universidad de Sonora Unidad Regional Sur, Apartado Postal, 85380, Navojoa, Sonora, Mexico

    Jesus A. Rosas-Rodríguez

  3. Departamento de Ciencias Químico-Biológicas, Universidad de Sonora, Apartado Postal, 83000, Hermosillo, Sonora, Mexico

    Aldo A. Arvizu-Flores

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  1. César Muñoz-Bacasehua
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  2. Jesus A. Rosas-Rodríguez
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  3. Aldo A. Arvizu-Flores
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  4. Elisa M. Valenzuela-Soto
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Correspondence to Elisa M. Valenzuela-Soto.

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Muñoz-Bacasehua, C., Rosas-Rodríguez, J.A., Arvizu-Flores, A.A. et al. Role of potassium levels in pkBADH heterogeneity of NAD+ binding site. J Bioenerg Biomembr 52, 61–70 (2020). https://doi.org/10.1007/s10863-020-09827-7

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  • DOI: https://doi.org/10.1007/s10863-020-09827-7

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