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Functional complementation of yeast cytosolic pyrophosphatase by bacterial and plant H -translocating pyrophosphatases

 

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dc.creator Losada Villasante, Manuel
dc.creator Pérez Castiñeira, José Román
dc.creator López Marqués, Rosa Laura
dc.creator Villalba Montoro, José Manuel
dc.creator Serrano Delgado, Aurelio
dc.date.accessioned 2015-06-25T10:02:58Z
dc.date.available 2015-06-25T10:02:58Z
dc.date.issued 2002
dc.identifier.issn 1091-6490 es
dc.identifier.issn 0027-8424 es
dc.identifier.uri http://hdl.handle.net/11441/26079
dc.description.abstract Two types of proteins that hydrolyze inorganic pyrophosphate (PPi), very different in both amino acid sequence and structure, have been characterized to date: soluble and membrane-bound proton-pumping pyrophosphatases (sPPases and H+-PPases, respectively). sPPases are ubiquitous proteins that hydrolyze PPi releasing heat, whereas H+-PPases, so far unidentified in animal and fungal cells, couple the energy of PPi hydrolysis to proton movement across biological membranes. The budding yeast Saccharomyces cerevisiae has two sPPases that are located in the cytosol and in the mitochondria. Previous attempts to knock out the gene coding for a cytosolic sPPase (IPP1) have been unsuccessful, thus suggesting that this protein is essential for growth. Here, we describe the generation of a conditional S. cerevisiae mutant (named YPC-1) whose functional IPP1 gene is under the control of a galactose-dependent promoter. Thus, YPC-1 cells become growth arrested in glucose but they regain the ability to grow on this carbon source when transformed with autonomous plasmids bearing diverse foreign H+-PPase genes under the control of a yeast constitutive promoter. The heterologously expressed H+-PPases are distributed among different yeast membranes, including the plasma membrane, functional complementation by these integral membrane proteins being consistently sensitive to external pH. These results demonstrate that hydrolysis of cytosolic PPi is essential for yeast growth and that this function is not substantially affected by the intrinsic characteristics of the PPase protein that accomplishes it. Moreover, this is, to our knowledge, the first direct evidence that H+-PPases can mediate net hydrolysis of PPi in vivo. YPC-1 mutant strain constitutes a convenient expression system to perform studies aimed at the elucidation of the structure–function relationships of this type of proton pumps. es
dc.format application/pdf es
dc.language.iso eng es
dc.publisher National Academy of Sciences es
dc.relation.ispartof Proceedings of the National Academy of Sciences of the United States of America, 99(25), 15914–15919 es
dc.rights Attribution-NonCommercial-NoDerivatives 4.0 Internacional *
dc.rights.uri http://creativecommons.org/licenses/by-nc-nd/4.0/ *
dc.title Functional complementation of yeast cytosolic pyrophosphatase by bacterial and plant H -translocating pyrophosphatases es
dc.type info:eu-repo/semantics/article es
dc.rights.accessrights info:eu-repo/semantics/openAccess es
dc.contributor.affiliation Universidad de Sevilla. Departamento de Bioquímica Vegetal y Biología Molecular es
dc.relation.publisherversion 10.1073/pnas.242625399 es
dc.identifier.idus https://idus.us.es/xmlui/handle/11441/26079
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