Characterization of cross-species transcription and splicing from Penicillium to Saccharomyces cerevisiae

Heterologous expression of eukaryotic gene clusters in yeast has been widely used for producing high-value chemicals and bioactive secondary metabolites. However, eukaryotic transcription cis-elements are still undercharacterized, and the cross-species expression mechanism remains poorly understood. Here we used the whole expression unit (including original promoter, terminator, and open reading frame with introns) of orotidine 5\u27-monophosphate decarboxylases from 14 Penicillium species as a showcase, and analyzed their cross-species expression in Saccharomyces cerevisiae. We found that pyrG promoters from the Penicillium species could drive URA3 expression in yeast, and that inefficient cross-species splicing of Penicillium introns might result in weak cross-species expression. Thus, this study demonstrates cross-species expression from Penicillium to yeast, and sheds light on the opportunities and challenges of cross-species expression of fungi expression units and gene clusters in yeast without refactoring for novel natural product discovery

Identification of manganese superoxide dismutase from Sphingobacterium sp. T2 as a novel bacterial enzyme for lignin oxidation

The valorisation of aromatic heteropolymer lignin is an important unsolved problem in the development of a biomass-based biorefinery, for which novel high-activity biocatalysts are needed. Sequencing of the genomic DNA of lignin-degrading bacterial strain Sphingobacterium sp. T2 revealed no matches to known lignin-degrading genes. Proteomic matches for two manganese superoxide dismutase proteins were found in partially purified extracellular fractions. Recombinant MnSOD1 and MnSOD2 were both found to show high activity for oxidation of Organosolv and Kraft lignin, and lignin model compounds, generating multiple oxidation products. Structure determination revealed that the products result from aryl-Cα and Cα-Cβ bond oxidative cleavage and O-demethylation. The crystal structure of MnSOD1 was determined to 1.35 Å resolution, revealing a typical MnSOD homodimer harbouring a 5-coordinate trigonal bipyramidal Mn(II) centre ligated by three His, one Asp and a water/hydroxide in each active site. We propose that the lignin oxidation reactivity of these enzymes is due to the production of hydroxyl radical, a highly reactive oxidant. This is the first demonstration that MnSOD is a microbial lignin-oxidising enzyme