Detailed search for protein kinase(s) involved in plasma membrane H+-ATPase activity regulation of yeast cells.

Pereira, Renata Rebeca; Castanheira, Diogo Dias; Teixeira, Janaina Aparecida; Bouillet, Leoneide Érica Maduro; Ribeiro, Erica Milena de Castro; Trópia, Maria José Magalhães; Alvarez, Florencia; Correa, Lygia Fátima da Mata; Mota, Bruno Eduardo Fernandes; Conceição, Luís Eduardo Fernandes Rodrigues da; Castro, Ieso de Miranda; Brandão, Rogélio Lopes

Detailed search for protein kinase(s) involved in plasma membrane H+-ATPase activity regulation of yeast cells.

Date

2015

Authors

Pereira, Renata Rebeca

Castanheira, Diogo Dias

Teixeira, Janaina Aparecida

Bouillet, Leoneide Érica Maduro

Ribeiro, Erica Milena de Castro

Trópia, Maria José Magalhães

Alvarez, Florencia

Correa, Lygia Fátima da Mata

Mota, Bruno Eduardo Fernandes

Conceição, Luís Eduardo Fernandes Rodrigues da

Abstract

This study displays a screening using yeast strains deficient in protein kinases known to exist in Saccharomyces cerevisiae. From 95 viable single mutants, 20 mutants appear to be affected in the glucose-induced extracellular acidification. The mutants that are unaffected in calcium signaling were tested for their sensitivity to hygromycin B. Furthermore, we verified whether the remaining mutants produced enzymes that are appropriately incorporated at plasma membrane. Finally, we measure the kinetic properties of the enzyme in purified plasma membranes from glucose-starved as well as glucose-fermenting cells. We confirmed the kinase Ptk2 involvement in H+−ATPase regulation (increase of affinity for ATP). However, the identification of the kinase(s) responsible for phosphorylation that leads to an increase in Vmax appears to be more complex. Complementary experiments were performed to check how those protein kinases could be related to the control of the plasma membrane H+−ATPase and/or the potential membrane. In summary, our results did not permit us to identify the protein kinase(s) involved in regulating the catalytic efficiency of the plasma membrane H+−ATPase. Therefore, our results indicate that the current regulatory model based on the phosphorylation of two different sites located in the C-terminus tail of the enzyme could be inappropriate.

Keywords

Glucose signaling, Yeast protein kinases

Citation

PEREIRA, R. R. et al. Detailed search for protein kinase(s) involved in plasma membrane H+-ATPase activity regulation of yeast cells. FEMS Yeast Research, v. 15, p. fov003-fov012, 2015. Disponível em: <https://academic.oup.com/femsyr/article-lookup/doi/10.1093/femsyr/fov003> . Acesso em: 16 jun. 2017.