Glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas

Glycosidases constitute a diverse group of enzymes that catalyze the hydrolysis of glycosidic bonds, thus releasing the sugar or modifying the structure of the original substrate. In addition to hydrolysis, some glycosidases are also capable of catalyzing transglycosylation reactions. In this proces...

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Autor principal: Baglioni, Micaela
Otros Autores: Mazzaferro, Laura S.
Formato: Tesis doctoral acceptedVersion
Lenguaje:Español
Publicado: Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica 2024
Materias:
Glicosidasa
Flavonoides
Biocatálisis
Biorrefinería
Ciencias de la vida
Acceso en línea:http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=posgraafa&cl=CL1&d=HWA_7844
https://repositoriouba.sisbi.uba.ar/gsdl/collect/posgraafa/index/assoc/HWA_7844.dir/7844.PDF
Aporte de:
Repositorio Digital de la Universidad de Buenos Aires (UBA) de Universidad de Buenos Aires
id I28-R145-HWA_7844
record_format dspace
institution Universidad de Buenos Aires
institution_str I-28
repository_str R-145
collection Repositorio Digital de la Universidad de Buenos Aires (UBA)
language Español
orig_language_str_mv spa
topic Glicosidasa
Flavonoides
Biocatálisis
Biorrefinería
Ciencias de la vida
spellingShingle Glicosidasa
Flavonoides
Biocatálisis
Biorrefinería
Ciencias de la vida
Baglioni, Micaela
Glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas
topic_facet Glicosidasa
Flavonoides
Biocatálisis
Biorrefinería
Ciencias de la vida
description Glycosidases constitute a diverse group of enzymes that catalyze the hydrolysis of glycosidic bonds, thus releasing the sugar or modifying the structure of the original substrate. In addition to hydrolysis, some glycosidases are also capable of catalyzing transglycosylation reactions. In this process, sugar is transferred from a sugar donor substrate to a different nucleophilic acceptor other than water, thereby forming a new glycosidic bond. Both activities have various biotechnological applications that are constantly expanding. In this work, we studied bacterial and fungal glycosidases using a biochemical approach combined with the exploration of genomes sequenced by our laboratory and available in public databases (genome mining). The bacterium Actinoplanes missouriensis 431T and the fungus Aspergillus alliaceus DSM 813 were the central microorganisms of the study. Genomic context analysis of the gene encoding ?R?G diglycosidase from A. missouriensis 431T (BAL86042.1) revealed the presence of an ?-L-rhamnosidase under the same transcriptional regulator. This genetic pattern was also identified in five strains of the Actinoplanes genus, allowing us to propose a catabolic pathway for hesperidin hydrolysis and subsequent utilization of the monosaccharides. In contrast, induction studies by flavonoids (naringin, rutin) in A. alliaceus DSM 813 revealed a preferential pathway of sequential deglycosylation involving an ?- rhamnosidase and ?-glucosidase. Recombinant enzymes and commercial enzymatic preparations were used in the synthesis of value-added compounds (hesperetin 7-O-glucoside, rutinose, 4-nitrophenyl ?-rutinoside, and glyceryl rutinoside). The compound synthesized by the ?-L-rhamnosidase of A. missourensis 431T, hesperetin 7-O-glucoside, was used in the study of regiospecificity of the ?R?G I diglycosidase from Acremonium sp. DSM 24697, allowing the detection for the first time of monoglucosidase activity of this enzyme. It is noteworthy that hesperetin 7-O-glucoside is the glucosidic derivative of the substrate with the highest specificity for ?R?G I, hesperidin. Subsequently, by engineering the reaction medium, the selectivity of the commercial enzymatic cocktail AromaseTM H2 glycosidase mixture was directed, allowing the development of selective processes for rutinose production and synthesis of 4-nitrophenol ?-rutinoside. Finally, a bioprocess was developed for the synthesis and purification of glyceryl rutinoside with the ?R?G I enzyme from Acremonium sp. DSM 24697. Glycerol and citrus fruit processing residue were used as glycosidic donor substrates, proposing an alternative in the biorefinery of these fruits.
author2 Mazzaferro, Laura S.
author_facet Mazzaferro, Laura S.
Baglioni, Micaela
format Tesis doctoral
Tesis doctoral
acceptedVersion
author Baglioni, Micaela
author_sort Baglioni, Micaela
title Glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas
title_short Glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas
title_full Glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas
title_fullStr Glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas
title_full_unstemmed Glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas
title_sort glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas
publisher Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica
publishDate 2024
url http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=posgraafa&cl=CL1&d=HWA_7844
https://repositoriouba.sisbi.uba.ar/gsdl/collect/posgraafa/index/assoc/HWA_7844.dir/7844.PDF
work_keys_str_mv AT baglionimicaela glicosidasascomoherramientasparaladiversificacionestructuraldemoleculasbioactivas
AT baglionimicaela glycosidasestoolboxforthestructuraldiversificationofbioactivecompounds
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spelling I28-R145-HWA_78442025-08-01 Glycosidases constitute a diverse group of enzymes that catalyze the hydrolysis of glycosidic bonds, thus releasing the sugar or modifying the structure of the original substrate. In addition to hydrolysis, some glycosidases are also capable of catalyzing transglycosylation reactions. In this process, sugar is transferred from a sugar donor substrate to a different nucleophilic acceptor other than water, thereby forming a new glycosidic bond. Both activities have various biotechnological applications that are constantly expanding. In this work, we studied bacterial and fungal glycosidases using a biochemical approach combined with the exploration of genomes sequenced by our laboratory and available in public databases (genome mining). The bacterium Actinoplanes missouriensis 431T and the fungus Aspergillus alliaceus DSM 813 were the central microorganisms of the study. Genomic context analysis of the gene encoding ?R?G diglycosidase from A. missouriensis 431T (BAL86042.1) revealed the presence of an ?-L-rhamnosidase under the same transcriptional regulator. This genetic pattern was also identified in five strains of the Actinoplanes genus, allowing us to propose a catabolic pathway for hesperidin hydrolysis and subsequent utilization of the monosaccharides. In contrast, induction studies by flavonoids (naringin, rutin) in A. alliaceus DSM 813 revealed a preferential pathway of sequential deglycosylation involving an ?- rhamnosidase and ?-glucosidase. Recombinant enzymes and commercial enzymatic preparations were used in the synthesis of value-added compounds (hesperetin 7-O-glucoside, rutinose, 4-nitrophenyl ?-rutinoside, and glyceryl rutinoside). The compound synthesized by the ?-L-rhamnosidase of A. missourensis 431T, hesperetin 7-O-glucoside, was used in the study of regiospecificity of the ?R?G I diglycosidase from Acremonium sp. DSM 24697, allowing the detection for the first time of monoglucosidase activity of this enzyme. It is noteworthy that hesperetin 7-O-glucoside is the glucosidic derivative of the substrate with the highest specificity for ?R?G I, hesperidin. Subsequently, by engineering the reaction medium, the selectivity of the commercial enzymatic cocktail AromaseTM H2 glycosidase mixture was directed, allowing the development of selective processes for rutinose production and synthesis of 4-nitrophenol ?-rutinoside. Finally, a bioprocess was developed for the synthesis and purification of glyceryl rutinoside with the ?R?G I enzyme from Acremonium sp. DSM 24697. Glycerol and citrus fruit processing residue were used as glycosidic donor substrates, proposing an alternative in the biorefinery of these fruits. Fil: Baglioni, Micaela. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Buenos Aires, Argentina Mazzaferro, Laura S. Miranda, María Victoria Breccia, Javier D. Baglioni, Micaela 2024-05-13 Las glicosidasas constituyen un grupo diverso de enzimas que catalizan la hidrólisis de las uniones glicosídicas, liberando así el azúcar o modificando la estructura del sustrato original. Además de la hidrólisis, algunas glicosidasas también son capaces de catalizar reacciones de transglicosilación. En este proceso, un azúcar es transferido desde un sustrato donante de azúcar a otro aceptor nucleofílico diferente del agua, formando así un nuevo enlace glicosídico. Ambas actividades tienen diversas aplicaciones biotecnológicas en constante expansión. En este trabajo estudiamos glicosidasas bacterianas y fúngicas mediante un enfoque bioquímico combinado con la exploración de genomas secuenciados por nuestro laboratorio y disponibles en bases de datos públicas (genome mining). La bacteria Actinoplanes missouriensis 431T y el hongo Aspergillus alliaceus DSM 813 fueron los microorganismos centrales del estudio. El análisis del contexto genómico del gen que codifica la diglicosidasa ?R?G de A. missouriensis 431T (BAL86042.1) revelo la presencia de una ?-L-ramnosidasa bajo el mismo regulador transcripcional. Este patrón genético se identificó también en cinco cepas del género Actinoplanes, lo que permitió proponer una vía catabólica para la hidrólisis de hesperidina y la utilización subsiguiente de los monosacáridos. En contraste, los estudios de producción de actividad glicosidasa en medios de cultivo con flavonoides (naringina, rutina) en A. alliaceus revelaron una vía preferencial de desglicosilación secuencial que involucra una ?- ramnosidasa y ?-glucosidasa. Se emplearon enzimas recombinantes y preparados enzimáticos comerciales en la síntesis de compuestos de valor agregado (hesperetin 7-O-glucósido, rutinosa, 4-nitrofenil ?-rutinósido y gliceril rutinósido). El compuesto sintetizado por la ?-L-ramnosidasa de A. missourensis 431T, hesperetin 7-O-glucósido, se empleó en el estudio de regioespecificidad de la diglicosidasa ?R?G I de Acremonium sp. DSM 24697 permitiendo detectar por primera vez actividad monoglucosidasa de esta enzima. Cabe destacar que hesperetin 7-O-glucósido es el derivado glucosídico del sustrato de mayor especificidad de la ?R?G I, hesperidina. Posteriormente, mediante ingeniería del medio de reacción se dirigió la selectividad de la mezcla de glicosidasas del cóctel enzimático comercial AromaseTM H2, permitiendo el desarrollo de procesos selectivos de producción de rutinosa y síntesis de 4-nitrofenol ?-rutinósido. Finalmente, se desarrolló un bioproceso para la síntesis y purificación de gliceril rutinósido con la enzima ?R?G I de Acremonium sp. DSM 24697. Se utilizaron los sustratos glicerol y el residuo del procesamiento de frutas cítricas como donante glicosídico, proponiendo una alternativa más en la biorrefinería de estas frutas. application/pdf Talou, Julián Rodríguez Stortz, Carlos Briand, Laura Estefanía Glicosidasa Flavonoides Biocatálisis Biorrefinería spa Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by-nc-nd/2.5/ar/ Ciencias de la vida Doctora de la Universidad de Buenos Aires en Ciencias Bioquímicas Glicosidasas como herramientas para la diversificación estructural de moléculas bioactivas Glycosidases toolbox for the structural diversification of bioactive compounds info:eu-repo/semantics/doctoralThesis info:ar-repo/semantics/tesis doctoral info:eu-repo/semantics/acceptedVersion http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=posgraafa&cl=CL1&d=HWA_7844 https://repositoriouba.sisbi.uba.ar/gsdl/collect/posgraafa/index/assoc/HWA_7844.dir/7844.PDF

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