Novel flavonoid C-8 hydroxylase from Rhodotorula glutinis: identification, characterization and substrate scope

• DOI: 10.1186/s12934-022-01899-x
• Publication Date: 2022-08-29T06:03:36Z
• Authors: Kinga Dulak; Sandra Sordon; Agata Matera; Bartosz Kozak; Ewa Huszcza; Jarosław Popłoński
• Keywords: Rhodotorula; Identification; Scope (computer science)

Abstract Background The regioselective hydroxylation of phenolic compounds, especially flavonoids, is still a bottleneck classical organic chemistry that could be solved using enzymes with high activity and specificity. Yeast Rhodotorula glutinis KCh735 in known to catalyze the C-8 flavones flavanones. enzyme F8H (flavonoid C8-hydroxylase) involved reaction, but specific gene has not yet been identified. In this work, we present identification, heterologous expression characterization first ortho -hydroxylase from yeast. Results Differential transcriptome analysis homology bacterial monooxygenases, including also FAD-dependent motif GD characteristic for flavin-dependent provided set coding sequences among which RgF8H was Phylogenetic suggests member flavin monooxygenase group active on flavonoid substrates. Analysis recombinant protein showed catalyzes C8-hydroxylation naringenin, hesperetin, eriodyctiol, pinocembrin, apigenin, luteolin, chrysin, diosmetin 7,4ʹ-dihydroxyflavone. presence C7-OH necessary enzymatic indicating -hydroxylation mechanism. requires NADPH coenzyme regeneration prosthetic group, displays very low hydroxyperoxyflavin decupling rate, addition FAD significantly increases its activity. Conclusions This study presents identification yeast hydroxylase responsible flavonoids (F8H). biochemically characterized applied vitro cascade Bacillus megaterium glucose dehydrogenase reactions. High vivo Escherichia coli enable further synthetic biology application towards production rare highly antioxidant compounds.