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Epistatic mutations in ISC metabolism synergize with cell cycle regulation and the PPP to enhance xylose fermentation and acetic acid tolerance in industrial yeast

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Autor(es):
dos Santos, Leandro Vieira ; Palermo, Gisele Cristina de Lima ; Costa, Paulo Emilio dos Santos ; Almeida, Ludimila Dias ; Carazzolle, Marcelo Falsarella ; Pereira, Goncalo Amarante Guimaraes
Número total de Autores: 6
Tipo de documento: Artigo Científico
Fonte: Bioresource Technology; v. 439, p. 15-pg., 2026-01-26.
Resumo

Efficient utilization of complex biomass-derived sugars and tolerance to inhibitors are key requirements for the viability of lignocellulosic-based biorefineries. In this study, a two-stage evolution of an industrial yeast strain engineered with a xylose isomerase pathway yielded strain AceY.14, which exhibited improved fermentative performance and increased tolerance to acetic acid. Whole-genome sequencing of the evolved strain identified SNPs in ZWF1, a component of the pentose phosphate pathway (PPP), and in the G1 cyclin gene CLN3, both of which were functionally validated through CRISPR and reverse engineering. The zwf1E191D mutation reduced xylitol accumulation, alleviating inhibition of xylose isomerase and enhancing flux through the non-oxidative branch of the PPP, while the frameshift cln3T556fs mutation unexpectedly improved acetic acid tolerance and xylose consumption in the evolved strain, also affecting cell size and growth. Genome sequencing of AceY.14 also revealed a significant reduction in the xylA gene copy number, likely decreasing the metabolic burden associated with high xylose isomerase expression. A synergistic effect was observed in the isu1 Delta/zwf1 Delta double mutant, further boosting xylose consumption rates. A diploid derivative (AceY-2n) demonstrated high productivity and robustness in fermentations using hydrolysates from various lignocellulosic feedstocks, highlighting the strain's potential for industrial-scale applications. These findings reveal novel metabolic targets for strain optimization and offer valuable insights for the rational engineering of yeast platforms for sustainable biofuel and bioproduct production. (AU)

Processo FAPESP: 20/07918-7 - Abordagens sistêmicas integradas para o desenvolvimento de leveduras como plataformas microbianas em biorrefinarias
Beneficiário:Leandro Vieira dos Santos
Modalidade de apoio: Auxílio à Pesquisa - Programa BIOEN - Regular
Processo FAPESP: 20/02936-7 - Mecanismos moleculares envolvidos na sinalização celular e regulação gênica da assimilação de xilose em linhagens modificadas de Saccharomyces cerevisiae
Beneficiário:Gisele Cristina de Lima Palermo
Modalidade de apoio: Bolsas no Brasil - Doutorado Direto
Processo FAPESP: 19/06942-4 - Elucidação das bases moleculares da fermentação de arabinose e xilose em linhagens de Saccharomyces cerevisiae
Beneficiário:Paulo Emílio dos Santos Costa
Modalidade de apoio: Bolsas no Brasil - Mestrado
Processo FAPESP: 17/08519-6 - Construção de um atlas genômico para engenharia racional de Saccharomyces cerevisiae visando a produção de etanol de segunda-geração
Beneficiário:Leandro Vieira dos Santos
Modalidade de apoio: Auxílio à Pesquisa - Regular