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bioRxiv · 10.1101/2023.05.10.540258

Influence of expression protocols on Gα biochemical activity: kinetics of plant and mammalian G protein cycles

Gookin, T. E.·Chakravorty, D.·Assmann, S. M.
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Abstract

Heterotrimeric G proteins are a class of signal transduction complexes with broad roles in human health and agriculturally important plant traits. In the classic paradigm, guanine nucleotide binding to the G subunit regulates the activation status of the complex. Using the Arabidopsis thaliana G subunit, GPA1, we developed a rapid StrepII-tag mediated purification method that facilitates isolation of protein with increased enzymatic activities as compared to conventional methods, and is demonstrably also applicable to mammalian G subunits. We subsequently utilized domain swaps of GPA1 and human GNAO1 to demonstrate the instability of recombinant GPA1 is a function of the interaction between the Ras and helical domains, and can be partially uncoupled from the rapid nucleotide binding kinetics displayed by GPA1.

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Gookin, T. E., Chakravorty, D., Assmann, S. M.. 2023-05-10. Influence of expression protocols on Gα biochemical activity: kinetics of plant and mammalian G protein cycles. https://doi.org/10.1101/2023.05.10.540258

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