English   español  
Please use this identifier to cite or link to this item: http://hdl.handle.net/10261/188516
logo share SHARE   Add this article to your Mendeley library MendeleyBASE
Visualizar otros formatos: MARC | Dublin Core | RDF | ORE | MODS | METS | DIDL | DATACITE
Exportar a otros formatos:


Engineering the unspecific peroxygenase, a wide reaction range biocatalyst, by directed evolution

AuthorsMolina-Espeja, Patricia ; González-Pérez, David ; Alcalde Galeote, Miguel ; García-Ruiz, Eva
Issue Date7-Apr-2014
Citation3rd Multistep Enzyme Catalyzed Processes Congress (2014)
AbstractThe unspecific peroxygenase (UPO) is a new type of heme-thiolate enzyme with an exclusive self-sufficient mono(per)oxygenase activity and many attractive applications in organic synthesis amongst other biotechnological uses (1, 2). In this work, the UPO1 gene from the basidiomiycete Agrocybe aegerita was subjected to directed evolution using Saccharomyces cerevisiae as heterologous host. To promote functional expression, several fusions were tested comprising different signal peptides attached to the mature protein. Over 9,000 clones were screened with an ad-hoc dual-colorimetric assay that permitted to assess both peroxidative (ABTS as substrate) and oxygen-transfer activities (NBD as substrate) (3). After five generations of evolution (including random, hybrid and rational approaches such as mutagenic PCR, MORPHING (4) and sitedirected mutagenesis, respectively), 9 mutations were introduced providing a total activity improvement of 3,250-fold without jeopardizing the protein stability. The evolved UPO1 was active and highly stable in the presence of extreme concentrations of organic cosolvents. Mutations at the hydrophobic core of the signal peptide enhanced secretion levels whereas some mutations placed in the neighborhood of the heme access
DescriptionTrabajo presentado en el 3rd Multistep Enzyme Catalyzed Processes Congress, celebrado en Madrid (España) del 07 al 10 de abril de 2014.
Appears in Collections:(ICP) Comunicaciones congresos
Files in This Item:
File Description SizeFormat 
accesoRestringido.pdf15,38 kBAdobe PDFThumbnail
Show full item record
Review this work

WARNING: Items in Digital.CSIC are protected by copyright, with all rights reserved, unless otherwise indicated.