Pre-development On Directional Modification Of Value-added Pentacyclic Meroditerpenoid Chevalone E By Chemo-Enzymatic Catalysis

Endophytic fungi of medicinal plants are an important resource of natural active substances.Meroterpenoids in fungal secondary metabolites have complex ring structure and have widely biological activities,such as anti-inflammatory,antiproliferative,antitumor,antibacterial,antifungal,anti-allergic,antioxidant,antihyperlipidemic,cytotoxic and other activities.Although meroterpenoids have high medicinal value,their natural expression yield is too low and the economic cost of acquiring them is too high.The economic cost of obtaining meroterpenoids from heterologous expression of Aspergillus oryzae is relatively low.Chemo-enzymatic catalysis modification is an important means of derivatization of compound structure,which happens to be a concept often used in drug design to appropriately change the structure of drug molecules to reduce toxic side effects of drugs.In this paper,with the help of heterologous expression of Aspergillus oryzae,we have found a gene cluster that can produce pentacyclic meroditerpenoids chevalone E(1)and its derivatives in Aspergillus versicolor 0312,and obtained a high-yielding compound 1 strain of Aspergillus oryzae.And then,the chemo-enzyme olcF’(short-chain dehydrogenase)of the olc’gene cluster of Aspergillus felis 0260 in endophytic fungi in Paris polyphylla var.yunnanensis,can modify the A ring of compound 1 to generate chevalone F(9),and then heterologously expressed in the Aspergillus oryzae host in combination with P450 oxidases cle2 and cle4 to produce 7new chevalone analogues(compounds 10-16).The new compound 10 contains a characteristic structure of a semiacetal bridge,which can be converted to spironolactone compound 17 by simple chemical synthesis.At the same time,compounds 9-12 have synergistic doxorubicin(DOX)antitumor activity.In this paper,the biosynthetic pathway of chevalone analogues was deduced by feeding experiments,enzyme reaction assays,and heterologous expression experiments of Aspergillus oryzae.Fortunately,it is the first time that meroditerpenoid compounds have been directional modified by chemo-enzymatic catalysis.It was revealed that the targeted chemo-enzyme(OlcF’protein)is a NAD~+-dependent short chain dehydrogenase.The important natural product of the endophytic fungy Aspergillus versicolor0312 cle gene cluster in Paris polyphylla var.yunnanensis was directionally modified by chemo-enzyme catalysis.And chevalone analogues with added value were obtained as chevalone E,which provided an idea for the discovery and construction of new natural products with synergistic activity.These strategies provided a certain reference for further transformation of scientific and technological achievements.