Supporting and additional data to FWF project P 32901-B "B-ring hydroxylation in the flavonoid pathway".More information can be found in the related journal articles (see the section "related works").

Project context

The project 'B-Ring hydroxylation in the flavonoid pathway' was designed as a follow up proposal to P29552-B29, which started the establishment of the first crystal structure of a cytochrome P450 dependent monooxygenase (CYP) of the flavonoid pathway using the specialized CYP75B member chalcone 3-hydroxylase (CH3H) as a model. CH3H is closely related to the prominent flavonoid 3'-hydroxylase (F3'H) and a key enzyme in the biosynthesis of anthochlor pigments, which provide yellow flower colour in a number of ornamental plants, form UV-honey guides in certain Asteraceae species and also show health-beneficial effects of chalcones in humans. The project aimed at enabling an understanding of the structure-function relationship of the CYPs determining the B-ring hydroxylation pattern of flavonoid structures (CH3H, F3'H, F3'5'H). The identification of the amino acids involved in substrate binding and their mode of action is of key interest as this allows targeted application in biotransformation, and breeding of plants with increased disease tolerance and/or improved aesthetic qualities. Procedures were established for the recombinant production of a soluble variant of CH3H and three cytochrome P450 reductases from different ornamental species with high yields, which allowed characterization of the purified enzyme and provided new insights into CH3H substrate specificity. Limited long-time stability of CH3H, however, so far impeded crystallization of the enzyme. Studies of F3'H from Malus sp. concentrated on the potential contribution to dihydrochalcone hydroxylation and its impact for watercore, an internal physiological disorder of apple. The results also contributed to the creation of the first genome edited poinsettias towards orange flower colour. For F3'5'H, amino acids essential for enzyme activity were identified for the first time. The project involved Heidi Halbwirth (expertise in flavonoid biosynthesis and hydroxylating enzymes), Oliver Spadiut (expertise in protein production and purification) and Christian Molitor (expertise in crystallization and modelling). This project led to seven publications in peer-reviewed journals.

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All data are licensed under the CC BY 4.0 license.

Funding

FWF (Austrian Science Fund) – B-ring hydroxylation in the flavonoid pathway (P32901-B)