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Thermal Stability of Dopamine Transporters

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Abstract

The thermal stabilities of the rat and mouse dopamine transporter (DAT) proteins were studied within the temperature range of 0–37 °C. The inactivation of the protein was followed by monitoring changes in radioligand-specific binding. We found that the process followed a rate equation with first-order kinetics and was characterized by having a single rate constant k inact. The activation energies (E a) that were calculated from the Arrhenius plots (ln k inact vs. 1/T) were 43 ± 5 and 45 ± 6 kJ/mol for the rat (rDAT) and mouse (mDAT) transporters, respectively, and 44 ± 7 kJ/mol for rDAT from PC-6.3 cell line. These E a values were similar to the E a values of thermal inactivation of the muscarinic receptor from rat brain cortex and to the thermal inactivation of other transmembrane proteins. However, all of these activation energy values were significantly lower than the E a values for soluble single-subunit proteins of similar size. These results therefore suggest that the thermal stability of transmembrane proteins may be governed to a significant extent by cell membrane properties and by interactions between the membrane components and the protein. In contrast, the stability of soluble proteins seems to be mostly governed by protein structure and size, which determine the sum of the stabilizing intramolecular interactions within the protein molecule. It is therefore not surprising that cell membrane properties and composition may have significant effects on the functional properties of transmembrane proteins.

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Acknowledgments

This work was funded by the Grant IUT20-15 of the Estonian Ministry of Education and Research. The authors are grateful to Dr. Monika Jürgenson from the Institute of Pharmacology, University of Tartu for providing samples of rodent striata. The authors thank Professor Mart Saarma and MSc Maria Lume for providing PC-6.3 cells.

Conflict of interest

The authors declare that there are no conflict of interest.

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Authors and Affiliations

  1. Chair of Organic Chemistry, Institute of Chemistry, University of Tartu, Ravila 14a, 50411, Tartu, Estonia

    Siim Kukk & Jaak Järv

  2. Center for Psychiatric Research, Department of Clinical Neuroscience, Karolinska Institute, Stockholm, Sweden

    Vladimir Stepanov

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  1. Siim Kukk
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  2. Vladimir Stepanov
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  3. Jaak Järv
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Correspondence to Siim Kukk.

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Kukk, S., Stepanov, V. & Järv, J. Thermal Stability of Dopamine Transporters. J Membrane Biol 248, 775–781 (2015). https://doi.org/10.1007/s00232-015-9794-9

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