RT Journal Article
T1 Secretion of Recombinant Pro- and Mature Fungal α-Sarcin Ribotoxin by the Methylotrophic YeastPichia pastoris:The Lys–Arg Motif Is Required for Maturation
A1 Martínez Ruiz, Antonio
A1 Martínez Del Pozo, Álvaro
A1 Lacadena García-Gallo, Francisco Javier
A1 Mancheño Gómez, José Miguel
A1 Oñaderra Sánchez, Mercedes
A1 Carlos López-Otı́n, 
A1 Gavilanes Franco, José Gregorio
AB α-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.
PB Elsevier
SN 1046-5928
YR 1998
FD 1998-04
LK https://hdl.handle.net/20.500.14352/103373
UL https://hdl.handle.net/20.500.14352/103373
LA eng
NO Martínez-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.
DS Docta Complutense
RD 28 jul 2024