RESUMO
Industrial production of semi-synthetic cephalosporins by Penicillium chrysogenum requires supplementation of the growth media with the side-chain precursor adipic acid. In glucose-limited chemostat cultures of P. chrysogenum, up to 88% of the consumed adipic acid was not recovered in cephalosporin-related products, but used as an additional carbon and energy source for growth. This low efficiency of side-chain precursor incorporation provides an economic incentive for studying and engineering the metabolism of adipic acid in P. chrysogenum. Chemostat-based transcriptome analysis in the presence and absence of adipic acid confirmed that adipic acid metabolism in this fungus occurs via ß-oxidation. A set of 52 adipate-responsive genes included six putative genes for acyl-CoA oxidases and dehydrogenases, enzymes responsible for the first step of ß-oxidation. Subcellular localization of the differentially expressed acyl-CoA oxidases and dehydrogenases revealed that the oxidases were exclusively targeted to peroxisomes, while the dehydrogenases were found either in peroxisomes or in mitochondria. Deletion of the genes encoding the peroxisomal acyl-CoA oxidase Pc20g01800 and the mitochondrial acyl-CoA dehydrogenase Pc20g07920 resulted in a 1.6- and 3.7-fold increase in the production of the semi-synthetic cephalosporin intermediate adipoyl-6-APA, respectively. The deletion strains also showed reduced adipate consumption compared to the reference strain, indicating that engineering of the first step of ß-oxidation successfully redirected a larger fraction of adipic acid towards cephalosporin biosynthesis.
Assuntos
Cefalosporinas/biossíntese , Engenharia Metabólica/métodos , Penicillium chrysogenum/metabolismo , Acil-CoA Desidrogenases/genética , Acil-CoA Desidrogenases/metabolismo , Acil-CoA Oxidase/genética , Acil-CoA Oxidase/metabolismo , Adipatos/metabolismo , Deleção de Genes , Regulação Fúngica da Expressão Gênica , Glucose/metabolismo , Mitocôndrias/enzimologia , Mitocôndrias/genética , Oxirredução , Peroxissomos/enzimologia , Peroxissomos/genética , TranscriptomaRESUMO
Diselenides are known precatalysts for catalytic oxidations. The use of these widely available compounds as catalysts for the oxidation of alcohols was investigated, still leaving many mechanistic questions to be answered. By using a range of analytical techniques, we found evidence for the unexpected involvement of seleninic anhydride in the catalytic mechanism. The activation time of the diselenide and its influence on the oxidation reaction itself was also investigated. On the basis of these findings, an improved protocol for the selective oxidation of activated alcohols was devised resulting in significantly decreased catalyst loadings (<1%).
Assuntos
Álcoois/química , Derivados de Benzeno/química , Compostos Organosselênicos/química , terc-Butil Hidroperóxido/química , Anidridos/química , Anisóis/química , Álcool Benzílico/química , Calorimetria , Catálise , Estrutura Molecular , OxirreduçãoRESUMO
[reaction: see text] Partially protected thioglycuronic acids are prepared efficiently by chemo- and regioselective oxidation of the corresponding thioglycosides using the TEMPO/BAIB reagent combination. After esterification, the thioglycuronic acids proved to be useful as both donor and acceptor in sulfonium-mediated condensations toward acidic di- and trisaccharides.