In Vitro Comparison of the Activity Requirements and Substrate Specificity of Human and Triboleum castaneum PINK1 Orthologues

PloS One
Liesbeth AertsVanessa A Morais

Abstract

Mutations in the gene encoding the mitochondrial kinase PINK1 cause early-onset familial Parkinson's disease. To understand the biological function of PINK1 and its role in the pathogenesis of Parkinson's disease, it is useful to study its kinase activity towards substrates both in vivo and in vitro. For in vitro kinase assays, a purified Triboleum castaneum PINK1 insect orthologue is often employed, because it displays higher levels of activity when compared to human PINK1. We show, however, that the activity requirements, and more importantly the substrate specificity, differ between both orthologues. While Triboleum castaneum PINK1 readily phosphorylates the PINKtide peptide and Histone H1 in vitro, neither of these non-physiological substrates is phosphorylated by human PINK1. Nonetheless, both Tc and human PINK1 phosphorylate Parkin and Ubiquitin, two physiological substrates of PINK1. Our results show that the substrate selectivity differs among PINK1 orthologues, an important consideration that should be taken into account when extrapolating findings back to human PINK1.

References

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Citations

May 18, 2020·Neuroscience Research·Edgar Djaha Yoboue, Enza Maria Valente
May 1, 2021·Life·Filipa Barroso Gonçalves, Vanessa Alexandra Morais

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Methods Mentioned

BETA
peptide library
PCR
restriction digest
electrophoresis
transfection
immunoprecipitation

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