Paasela, T. | bioRxiv Science

bioRxiv2025

A glycerol oxidase from Norway spruce (Picea abies L. Karst.) expands biochemical and structural attributes of the GMC oxidoreductase superfamily

O_LIInvestigations to uncover the versatility of carbohydrate active enzymes belonging to the auxiliary activity family 3 (AA3) have focused on microbial enzymes. Plant genomes also harbor AA3 encoding genes, and reverse genetics approaches have shown their importance in plant development. To date, however, detailed biochemical characterizations and structures of plant AA3 enzymes have not been reported. C_LIO_LIHere, we describe screening AA3 encoding genes from Norway spruce and the first in-depth biochemical and structural characterization of a plant AA3 oxidase, PaAOX1. PaAOX1 was crystallized, and the structure was solved by X-ray crystallography. C_LIO_LIPaAOX1 demonstrated highest oxidase activity against glycerol. It oxidized glycerol to glyceraldehyde, then further to glyceric acid, and showed preference for L-glyceraldehyde over D-glyceraldehyde. Despite the low sequence similarity with fungal alcohol oxidases, it had all the structural features of the typical AA3 enzyme. C_LIO_LIPaAOX1 has a novel activity among the characterized AA3s, and it showed a new structural arrangement of the region surrounding the catalytic center. Our findings contribute to a deeper understanding of the structural and functional aspects of enzymes in the GMC superfamily, and especially of alcohol oxidases, expanding the knowledge from fungi to plants. C_LI

biochemistry↗

bioRxiv2023

Transcriptome analysis reveals novel regulators of the Scots pine stilbene pathway

Stilbenes are developmentally induced metabolites in Scots pine heartwood where they have important role in protecting wood from decaying fungi. The stilbene pathway is also stress inducible, and ultraviolet (UV)-C radiation was among the first discovered artificial stress activators of the pathway. Many conditions that activate the pathway are known, but the specific transcriptional regulators and the biosynthetic enzyme responsible for activating the stilbene precursor cinnamate in the pathway are unknown. Here we describe the first large-scale transcriptomic analysis of pine needles in response to UV-C exposure and treatment with translational inhibitor cycloheximide, both activating the transcription of stilbene pathway genes, to uncover the candidates for the transcriptional regulation of the pathway and the cinnamate activating CoA ligase. We show that the regulation of pine stilbene pathway has shared features with grapevine stilbene pathway. In both species, the stilbene pathway was transcriptionally activated after UV-C treatment, protein phosphatase inhibitors and plant hormones ethylene and jasmonate. The pine stilbene synthase promoter retains its inducibility when transformed in plants that normally do not synthesize stilbenes. Pine stilbene synthase promoter was able to activate the expression of a reporter gene in Arabidopsis in response to UV-C exposure and phosphatase inhibitors, but not as response to plant hormone treatment. This indicates that stilbene synthase gene regulation occurs both via ancient stress-response pathway(s) but also via species specific regulators. With transcriptomic approach we identified candidate enzymes for cinnamate acting CoA ligase and transcription factors regulating the pathway.

plant biology↗