Molecular cloning of the plasma membrane H(+)-ATPase from Kluyveromyces lactis: a single nucleotide substitution in the gene confers ethidium bromide resistance and deficiency in K+ uptake.

AUTOR(ES)

Miranda, M

RESUMO

A Kluyveromyces lactis strain resistant to ethidium bromide and deficient in potassium uptake was isolated. Studies on the proton-pumping activity of the mutant strain showed that a decreased H(+)-ATPase specific activity was responsible for the observed phenotypes. The putative K. lactis PMA1 gene encoding the plasma membrane H(+)-ATPase was cloned by its ability to relieve the potassium transport defect of this mutant and by reversing its resistance to ethidium bromide. Its deduced amino acid sequence predicts a protein 899 residues long that is structurally colinear in its full length to H(+)-ATPases cloned from different yeasts, except for the presence of a variable N-terminal domain. By PCR-mediated amplification, we identified a transition from G to A that rendered the substitution of the fully conserved methionine at position 699 by isoleucine. We attribute to this amino acid change the low capacity of the mutant H(+)-ATPase to pump out protons.

Documentos Relacionados

• Trehalose-Mediated Inhibition of the Plasma Membrane H+-ATPase from Kluyveromyces lactis: Dependence on Viscosity and Temperature
• Role of the Plasma Membrane H+-ATPase in K+ Transport.
• Plasma Membrane H+-ATPase in Maize Roots Induced for NO3- Uptake.
• Cloning and characterization of the plasma membrane H(+)-ATPase from Candida albicans.
• Targeting of two Arabidopsis H(+)-ATPase isoforms to the plasma membrane.