PXD053024

PXD053024 is an original dataset announced via ProteomeXchange.

Title	Verazine Biosynthesis from Simple Sugars in Engineered Saccharomyces cerevisiae
Description	Steroidal alkaloids are FDA-approved drugs (e.g., Zytiga) and promising drug candidates/leads (e.g., cyclopamine); yet many of the ≥ 697 known steroidal alkaloid natural products remain underutilized as drugs because it can be challenging to scale their biosynthesis in their producing organisms. Cyclopamine is a steroidal alkaloid produced by corn lily (Veratrum spp.) plants, and it is an inhibitor of the Hedgehog (Hh) signaling pathway. Therefore, cyclopamine is an important drug candidate/lead to treat human diseases that are associated with dysregulated Hh signaling, such as basal cell carcinoma and acute myeloid leukemia. Cyclopamine and its semi-synthetic derivatives have been studied in (pre)clinical trials as Hh inhibitor-based drugs. However, challenges in scaling the production of cyclopamine have slowed efforts to improve its 1 efficacy and safety profile through (bio)synthetic derivatization, often limiting drug development to synthetic analogs of cyclopamine such as the FDA-approved drugs Odomzo, Daurismo, and Erivedge. If a platform for the scalable and sustainable production of cyclopamine were established, then its (bio)synthetic derivatization, clinical development, and, ultimately, widespread distribution could be accelerated. Ongoing efforts to achieve this goal include the biosynthesis of cyclopamine in Veratrum plant cell culture and the semi-/total chemical synthesis of cyclopamine. Herein, this work advances efforts towards a promising future approach: the biosynthesis of cyclopamine in engineered microorganisms. We completed the heterologous microbial production of verazine (biosynthetic precursor to cyclopamine) from simple sugars (i.e., glucose and galactose) in engineered Saccharomyces cerevisiae (S. cerevisiae) through the inducible upregulation of the native yeast mevalonate and lanosterol biosynthetic pathways, diversion of biosynthetic flux from ergosterol (i.e., native sterol in S. cerevisiae) to cholesterol (i.e., biosynthetic precursor to verazine), and expression of a refactored five-step verazine biosynthetic pathway containing eight heterologous enzymes sourced from seven different species. Importantly, S. cerevisiae-produced verazine was indistinguishable via liquid chromatography-mass spectrometry from both a commercial standard (Veratrum spp. plant-produced) and Nicotiana benthamiana-produced verazine. To the best of our knowledge, this is the first report describing the heterologous production of a steroidal alkaloid in an engineered yeast. Verazine production was increased through design-build-test-learn cycles to a final titer of 27 ± 2 μg/L (1.7 ± 0.1 μg/g DCW). Together, this research lays the groundwork for future microbial biosynthesis of cyclopamine, (bio)synthetic derivatives of cyclopamine, and other steroidal alkaloid natural products.
HostingRepository	PRIDE
AnnounceDate	2025-03-08
AnnouncementXML	Submission_2025-03-07_20:08:00.214.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Christopher Petzold
SpeciesList	scientific name: Saccharomyces cerevisiae (Baker's yeast); NCBI TaxID: 4932;
ModificationList	monohydroxylated residue; deamidated residue; iodoacetamide derivatized residue
Instrument	Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2024-06-11 14:53:28	ID requested
⏵ 1	2025-03-07 20:08:01	announced

Winegar PH, Hudson GA, Dell LB, Astolfi MCT, Reed J, Payet RD, Ombredane HCJ, Iavarone AT, Chen Y, Gin JW, Petzold CJ, Osbourn AE, Keasling JD, Verazine biosynthesis from simple sugars in engineered Saccharomyces cerevisiae. Metab Eng, 85():145-158(2024) [pubmed]

10.1016/j.ymben.2024.07.011;

submitter keyword: Verazine

Steroidal Alkaloid

Natural Product

Microbial Biosynthesis

Metabolic Engineering

Synthetic Biology

Christopher J. Petzold
contact affiliation	Staff Scientist Biological Systems & Engineering Division Lawrence Berkeley National Laboratory Berkeley CA 94720
contact email	cjpetzold@lbl.gov
lab head
Christopher Petzold
contact affiliation	Lawrence Berkeley National Laboratory
contact email	cjpetzold@lbl.gov
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD053024
     1. Label: PRIDE project
     2. Name: Verazine Biosynthesis from Simple Sugars in Engineered Saccharomyces cerevisiae