Descripción del proyecto
Secuencia y función proteínicas de enzimas no caracterizadas en hongos
Los rápidos progresos en la tecnología de secuenciación del genoma dejan aproximadamente el 30 % de las proteínas predichas como «huérfanas», es decir, no existe información sobre su función o sobre enzimas estrechamente emparentadas. Los hongos son productores prolíficos de metabolitos secundarios. Sin embargo, el estudio de los grupos de genes biosintéticos subyacentes todavía está dando sus primeros pasos. El objetivo del proyecto financiado con fondos europeos fORPHAN es desarrollar un flujo de trabajo de anotación de proteínas basado en la caracterización de enzimas del reino fúngico. Los investigadores proponen el uso de búsquedas en bases de datos de secuenciación profunda dirigida para identificar enzimas biosintéticas clave en genomas de hongos disponibles al público. Los mejores genes candidato se expresarán «in vitro» y en hospedadores microbianos recombinantes para caracterizar su actividad en diferentes sustratos mediante cristalografía de rayos X.
Objetivo
Fast advances in genome sequencing and sequence processing technology leave ~30% of predicted proteins as orphans, meaning without known function or closely related enzymes. Being able to assign a function to such orphans opens avenues to select for and design powerful biocatalysts – individual enzymes, biosynthetic pathways or entire organisms. The herein proposed research aims at developing a de-orphanizing pipeline “fORPHAN” based on computational and experimental characterization of enzymes from the fungal kingdom. Fungi are known to be prolific producers of secondary metabolites, however, the study of the underlying biosynthetic gene clusters is still a fairly young field. The use of automated search and annotation pipelines is currently limited by the small number of experimentally characterized genes and gene clusters that can be used to train the algorithms. Therefore, I propose to use deep targeted database searches to identify key biosynthetic enzymes in the publicly available genomes of fungi. First targets will be the thus-far uncharacterized family of putative chalcone isomerases and the recently discovered family of type III polyketide synthases, which both have great potential for in vivo and in vitro applications. Several gene candidates will be expressed in vitro and in recombinant microbial hosts, and tested for activity on a range of substrates. Interesting candidates will also be characterized by X-ray crystallography. This research will not only yield biotechnologically-relevant catalysts but also provide the bioinformatic foundation for more challenging genome mining projects.
Ámbito científico
- natural sciencesearth and related environmental sciencesgeologymineralogycrystallography
- natural sciencesbiological sciencesmicrobiologymycology
- natural scienceschemical sciencescatalysisbiocatalysis
- natural sciencesbiological sciencesgeneticsgenomes
- natural sciencesbiological sciencesbiochemistrybiomoleculesproteinsenzymes
Palabras clave
Programa(s)
Régimen de financiación
MSCA-IF - Marie Skłodowska-Curie Individual Fellowships (IF)Coordinador
9712CP Groningen
Países Bajos