Lpx1p SERIN PROTEASE PARTICIPATES IN THE GLUCOSE-INDUCED S. cerevisiae H +-ATPASE ACTIVATION

CAMPETELLI A N; ORTIZ E; MONESTEROLO N E; AMAIDEN M R; PREVITALI G; VALDEZ-TAUBAS J; CASALE C H

Puerto Madryn

Congreso; Sociedad Argentina de investigación Bioquímica y Biología Molecular; 2010

SAIB

We have shown that tubulin, in its acetylated form, interacts with some PATPases inhibiting their enzymatic activities. In S. cerevisiae, the plasma membrane H+-ATPase is inhibited by tubulin and, upon glucose addition, the ATPase is activated and the tubulin is degraded. Recently, through an exhaustive biochemical and physicochemical characterization we have shown that in S. cerevisiae there is a citosolic protease that interacts with tubulin, it has a MW of ~45 kDa, a pI of 8-9 and is inhibited by serine protease inhibitors. To identify this protease we performed a genetic screening of haploid knock out yeast strains loosing the ORFs corresponding to several proteases. The screening was based in the identification, after glucose treatment, of yeast strains with an altered H+ATPase activation and tubulin degradation phenotype. The strain YOR084, loosing the LPX1 gene, which codifies a serine protease of 44 kDa and pI 8.4 was deficient in H+-ATPase activation, at least by tubulin degradation when stimulated with glucose. When this KO strain was complemented with a vector carrying the wt copy of the LPX1 gene, the phenotype was rescued, indicating that this protease is involved in the H+-ATPase activation mechanism. Even though some authors have already proposed a possible involvement of a protease in this activation mechanism, it is the first report where the protease involvement is confirmed and the enzyme is identified.