Secretion of Recombinant Pro- and Mature Fungal α-Sarcin Ribotoxin by the Methylotrophic YeastPichia pastoris:The Lys–Arg Motif Is Required for Maturation

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dc.contributor.authorMartínez Ruiz, Antonio
dc.contributor.authorMartínez Del Pozo, Álvaro
dc.contributor.authorLacadena García-Gallo, Francisco Javier
dc.contributor.authorMancheño Gómez, José Miguel
dc.contributor.authorOñaderra Sánchez, Mercedes
dc.contributor.authorCarlos López-Otı́n
dc.contributor.authorGavilanes Franco, José Gregorio
dc.date.accessioned2024-04-23T11:01:57Z
dc.date.available2024-04-23T11:01:57Z
dc.date.issued1998-04
dc.description.abstractα-Sarcin is a ribosome-inactivating protein from the moldAspergillus giganteus.The methylotrophic yeastPichia pastorishas been transformed with two plasmids (pHILD2preαS and pHILS1preαS), which contain the complete α-sarcin cDNA, including its original fungal leader peptide, under the control of yeast alcohol oxidase promoter. The second one is indeed fused to the signal sequence ofP. pastorisacid phosphatase. The transformed yeasts secreted both mature and pro-α-sarcin. The presence of this pro-α-sarcin in the yeast extracellular medium is due to an inefficient recognition of the pro-sequence by a putative Kex2p-like endopeptidase. A third plasmid accounting for a single mutation of the α-sarcin leader peptide was designed to produce a more efficient Kex2p recognition motif. This approach resulted in the extracellular production of only the mature protein, suggesting the existence of a two-step mechanism for processing its leader peptide. This recombinant α-sarcin is identical to the original fungal protein, according to activity and spectroscopic criteria. In addition, pro-α-sarcin, which has been characterized for the first time, also exhibits ribonucleolytic activity as the mature protein does. Therefore, protection of the producing cells against this kind of ribotoxins may depend on an efficient recognition of the signal sequence followed by translocation of the nascent polypeptide to the endoplasmic reticulum.
dc.description.departmentDepto. de Bioquímica y Biología Molecular
dc.description.facultyFac. de Ciencias Químicas
dc.description.refereedTRUE
dc.description.statuspub
dc.identifier.citationMartínez-Ruiz A, Del Pozo AM, Lacadena J, Mancheño JM, Oñaderra M, López-Otín C, et al. Secretion of recombinant pro- and mature fungal α-sarcin ribotoxin by the methylotrophic yeast Pichia pastoris: The Lys-Arg motif is required for maturation. Protein Expression and Purification. 1998;12(3):315-22.
dc.identifier.doi10.1006/prep.1997.0846
dc.identifier.issn1046-5928
dc.identifier.urihttps://hdl.handle.net/20.500.14352/103373
dc.issue.number3
dc.journal.titleProtein Expression and Purification
dc.language.isoeng
dc.page.final322
dc.page.initial315
dc.publisherElsevier
dc.rights.accessRightsopen access
dc.subject.cdu577.1
dc.subject.ucmCiencias
dc.subject.unesco2302 Bioquímica
dc.titleSecretion of Recombinant Pro- and Mature Fungal α-Sarcin Ribotoxin by the Methylotrophic YeastPichia pastoris:The Lys–Arg Motif Is Required for Maturation
dc.typejournal article
dc.volume.number12
dspace.entity.typePublication
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