Biosynthesis of a ganoderic acid in Saccharomyces cerevisiae by expressing a cytochrome P450 gene from Ganoderma lucidum

다국어 초록 (Multilingual Abstract)

Ganoderic acid (GA), a triterpenoid from the traditional Chinese medicinal higher fungus Ganoderma lucidum, possesses antitumor and other significant pharmacological activities. Owing to the notorious difficulty and immaturity in genetic manipulation of higher fungi as well as their slow growth, biosynthesis of GAs in a heterologous host is an attractive alternative for their efficient bioproduction. In this study, using Saccharomyces cerevisiae as a host, we did a systematic screening of cytochrome P450 monooxygenase (CYP450) gene candidates from G. lucidum, which may be responsible for the GA biosynthesis from lanosterol but have not been functionally characterized. As a result, overexpression of a CYP450 gene cyp5150l8 was firstly found to produce an antitumor GA, 3‐hydroxy‐lanosta‐8, 24‐dien‐26 oic acid (HLDOA) in S. cerevisiae, as confirmed by HPLC, LC‐MS and NMR. A final titer of 14.5 mg/L of HLDOA was obtained at 120 hr of the yeast fermentation. Furthermore, our in vitro enzymatic experiments indicate that CYP5150L8 catalyzes a three‐step biotransformation of lanosterol at C‐26 to synthesize HLDOA. To our knowledge, this is the first report on the heterologous biosynthesis of GAs. The results will be helpful to the GA biosynthetic pathway elucidation and to future optimization of heterologous cell factories for GA production.
Biosynthesis from lanosterol to ganoderic acids (bioactive triterpenoids) has been an unresolved puzzling question for several decades, and discovery of the functional genes remains extremely challenging. The SJTU group did a systematic screening of CYP450 genes from traditional Chinese medicinal mushroom Ganoderma lucidum, and a gene responsible for converting lanosterol into a ganoderic acid was for the first time discovered. The CYP5150L8 catalyzed three‐step oxidations of lanosterol to 3‐hydroxy‐lanosta‐8, 24‐dien‐26 oic acid (HLDOA) in a synthetic biology chassis Saccharomyces cerevisiae.