Psychoactive Plant Database - Neuroactive Phytochemical Collection

1. Appl Microbiol Biotechnol. 2024 May 7;108(1):323. doi: 
10.1007/s00253-024-13157-8.

An unexpected role of EasD(af): catalyzing the conversion of chanoclavine 
aldehyde to chanoclavine acid.

Yu ZP(1)(2), An C(3), Yao Y(4), Yan JZ(5), Gao SS(5), Gu YC(6)(7), Wang 
CY(8)(9)(10), Cui C(11).

Author information:
(1)Key Laboratory of Marine Drugs, The Ministry of Education of China, Institute 
of Evolution & Marine Biodiversity, School of Medicine and Pharmacy, Ocean 
University of China, Qingdao, 266003, People's Republic of China.
(2)Beijing Institute for Drug Control, NMPA Key Laboratory for Research and 
Evaluation of Generic Drugs, Beijing Key Laboratory of Analysis and Evaluation 
on Chinese Medicine, Beijing, 102206, People's Republic of China.
(3)Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 
Tianjin, 300308, People's Republic of China.
(4)State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy 
of Sciences, Beijing, 100101, People's Republic of China.
(5)Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for 
Marine Science and Technology, Qingdao, 266003, People's Republic of China.
(6)Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for 
Marine Science and Technology, Qingdao, 266003, People's Republic of China. 
yucheng.gu@syngenta.com.
(7)Syngenta Jealott's Hill International Research Centre, Bracknell, Berkshire, 
RG42 6EY, UK. yucheng.gu@syngenta.com.
(8)Key Laboratory of Marine Drugs, The Ministry of Education of China, Institute 
of Evolution & Marine Biodiversity, School of Medicine and Pharmacy, Ocean 
University of China, Qingdao, 266003, People's Republic of China. 
changyun@ouc.edu.cn.
(9)Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for 
Marine Science and Technology, Qingdao, 266003, People's Republic of China. 
changyun@ouc.edu.cn.
(10)Beijing Institute for Drug Control, NMPA Key Laboratory for Research and 
Evaluation of Generic Drugs, Beijing Key Laboratory of Analysis and Evaluation 
on Chinese Medicine, Beijing, 102206, People's Republic of China. 
changyun@ouc.edu.cn.
(11)Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for 
Marine Science and Technology, Qingdao, 266003, People's Republic of China. 
cuichs@tib.cas.cn.

Ergot alkaloids (EAs) are a diverse group of indole alkaloids known for their 
complex structures, significant pharmacological effects, and toxicity to plants. 
The biosynthesis of these compounds begins with chanoclavine-I aldehyde (CC 
aldehyde, 2), an important intermediate produced by the enzyme EasDaf or its 
counterpart FgaDH from chanoclavine-I (CC, 1). However, how CC aldehyde 2 is 
converted to chanoclavine-I acid (CC acid, 3), first isolated from Ipomoea 
violacea several decades ago, is still unclear. In this study, we provide in 
vitro biochemical evidence showing that EasDaf not only converts CC 1 to CC 
aldehyde 2 but also directly transforms CC 1 into CC acid 3 through two 
sequential oxidations. Molecular docking and site-directed mutagenesis 
experiments confirmed the crucial role of two amino acids, Y166 and S153, within 
the active site, which suggests that Y166 acts as a general base for hydride 
transfer, while S153 facilitates proton transfer, thereby increasing the acidity 
of the reaction. KEY POINTS: • EAs possess complicated skeletons and are widely 
used in several clinical diseases • EasDaf belongs to the short-chain 
dehydrogenases/reductases (SDRs) and converted CC or CC aldehyde to CC acid • 
The catalytic mechanism of EasDaf for dehydrogenation was analyzed by molecular 
docking and site mutations.

© 2024. The Author(s).

DOI: 10.1007/s00253-024-13157-8
PMCID: PMC11076337
PMID: 38713233 [Indexed for MEDLINE]

Conflict of interest statement: The authors declare no competing interests.


2. Appl Environ Microbiol. 2023 Aug 30;89(8):e0079323. doi: 10.1128/aem.00793-23.
 Epub 2023 Jul 11.

Two Satellite Gene Clusters Enhance Ergot Alkaloid Biosynthesis Capacity of 
Aspergillus leporis.

Davis KA(1), Jones AM(1), Panaccione DG(1).

Author information:
(1)Division of Plant and Soil Sciences, West Virginia University, Morgantown, 
West Virginia, USA.

Ergot alkaloids are fungal specialized metabolites that are important in 
agriculture and serve as sources of several pharmaceuticals. Aspergillus leporis 
is a soil saprotroph that possesses two ergot alkaloid biosynthetic gene 
clusters encoding lysergic acid amide production. We identified two additional, 
partial biosynthetic gene clusters within the A. leporis genome containing some 
of the ergot alkaloid synthesis (eas) genes required to make two groups of 
clavine ergot alkaloids, fumigaclavines and rugulovasines. Clavines possess 
unique biological properties compared to lysergic acid derivatives. 
Bioinformatic analyses indicated the fumigaclavine cluster contained functional 
copies of easA, easG, easD, easM, and easN. Genes resembling easQ and easH, 
which are required for rugulovasine production, were identified in a separate 
gene cluster. The pathways encoded by these partial, or satellite, clusters 
would require intermediates from the previously described lysergic acid amide 
pathway to synthesize a product. Chemical analyses of A. leporis cultures 
revealed the presence of fumigaclavine A. However, rugulovasine was only 
detected in a single sample, prompting a heterologous expression approach to 
confirm functionality of easQ and easH. An easA knockout strain of Metarhizium 
brunneum, which accumulates the rugulovasine precursor chanoclavine-I aldehyde, 
was chosen as expression host. Strains of M. brunneum expressing easQ and easH 
from A. leporis accumulated rugulovasine as demonstrated through mass 
spectrometry analysis. These data indicate that A. leporis is exceptional among 
fungi in having the capacity to synthesize products from three branches of the 
ergot alkaloid pathway and for utilizing an unusual satellite cluster approach 
to achieve that outcome. IMPORTANCE Ergot alkaloids are chemicals produced by 
several species of fungi and are notable for their impacts on agriculture and 
medicine. The ability to make ergot alkaloids is typically encoded by a 
clustered set of genes that are physically adjacent on a chromosome. Different 
ergot alkaloid classes are formed via branching of a complex pathway that begins 
with a core set of the same five genes. Most ergot alkaloid-producing fungi have 
a single cluster of genes that is complete, or self-sufficient, and produce 
ergot alkaloids from one or occasionally two branches from that single cluster. 
Our data show that Aspergillus leporis is exceptional in having the genetic 
capacity to make products from three pathway branches. Moreover, it uses a 
satellite cluster approach, in which gene products of partial clusters rely on 
supplementation with a chemical intermediate produced via another gene cluster, 
to diversify its biosynthetic potential without duplicating all the steps.

DOI: 10.1128/aem.00793-23
PMCID: PMC10467348
PMID: 37432119 [Indexed for MEDLINE]

Conflict of interest statement: The authors declare no conflict of interest.


3. ACS Synth Biol. 2023 Apr 21;12(4):1133-1145. doi: 10.1021/acssynbio.2c00626. 
Epub 2023 Mar 29.

Modular Pathway Compartmentalization for Agroclavine Overproduction in 
Saccharomyces cerevisiae.

Wu N(1)(2), Yao M(1)(2), Xiao W(1)(2), Dong T(1)(2), Ma H(1)(2), Du X(3), Wang 
Y(1)(2), Yuan Y(1)(2).

Author information:
(1)Frontier Science Center for Synthetic Biology and Key Laboratory of Systems 
Bioengineering (Ministry of Education), School of Chemical Engineering and 
Technology, Tianjin University, Tianjin 300072, China.
(2)Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin 
University, Tianjin 300072, China.
(3)State Key Laboratory of Toxicology and Medical Countermeasures, Beijing 
Institute of Pharmacology and Toxicology, 100850 Beijing, China.

Agroclavine, which has anti-depressant activity and anti-Alzheimer effects, is 
the raw material used to synthesize ergo-based drugs. Although the production of 
agroclavine from Saccharomyces cerevisiae is possible, its yield is 
exceptionally low. The current study proposes a modular compartmentalization 
strategy for identifying and modifying the bottleneck step in agroclavine 
overproduction. The agroclavine synthetic pathway was reconstituted in yeast, 
and the best combination of Claviceps fusiformis EasA with Claviceps purpurea 
EasD/EasG was identified. According to the data on the expression and 
subcellular localization of agroclavine pathway proteins, the whole pathway was 
divided into two modules by chanoclavine-I. Separate enzyme distribution within 
the downstream module and low expression of DmaW and EasE in the upstream module 
were identified as the bottleneck steps in the pathway. The pathway efficiency 
was enhanced 2.06-fold when the downstream module was entirely anchored to the 
endoplasmic reticulum compartment. Increasing NADPH supply by overexpressing 
POS5 further improved the agroclavine yield by 27.4%. Altering the intracellular 
localization of DmaW from the peroxisome to the endoplasmic reticulum (ER) not 
only improved protein expression but also accelerated the accumulation of 
agroclavine by 59.9%. Integration of all modified modules into the host 
chromosome resulted in an improved yield of agroclavine at 101.6 mg/L with flask 
fermentation (a 241-fold improvement over the initial strain) and ultimately 
produced 152.8 mg/L of agroclavine on fed-batch fermentation. The current study 
unlocked the potential of S. cerevisiae in the advanced biosynthesis of ergot 
alkaloids. It also provides a promising strategy to reconstitute 
compartmentalized pathways.

DOI: 10.1021/acssynbio.2c00626
PMID: 36987837 [Indexed for MEDLINE]


4. Front Bioeng Biotechnol. 2022 Dec 21;10:1095464. doi: 
10.3389/fbioe.2022.1095464. eCollection 2022.

A hybrid system for the overproduction of complex ergot alkaloid chanoclavine.

Ma Y(1)(2)(3), Yan J(2), Yang L(2), Yao Y(1), Wang L(1), Gao SS(2)(4), Cui 
C(2)(4).

Author information:
(1)CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, 
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese 
Academy of Sciences, Beijing, China.
(2)Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 
Tianjin, China.
(3)University of Chinese Academy of Sciences, Beijing, China.
(4)National Technology Innovation Center of Synthetic Biology, Tianjin, China.

Synthetic biology-based methods (Sbio) and chemical synthesis (Csyn) are two 
independent approaches that are both widely used for synthesizing biomolecules. 
In the current study, two systems were combined for the overproduction of 
chanoclavine (CC), a structurally complex ergot alkaloid. The whole synthetic 
pathway for CC was split into three sections: enzymatic synthesis of 4-Br-Trp 
(4-Bromo-trptophan) using cell-lysate catalysis (CLC), chemical synthesis of 
prechanoclavine (PCC) from 4-Br-Trp, and overproduction CC from PCC using a 
whole-cell catalysis (WCC) platform. The final titer of the CC is over 3 g/L in 
this Sbio-Csyn hybrid system, the highest yield reported so far, to the best of 
our knowledge. The development of such a combined route could potentially avoid 
the limitations of both Sbio and Csyn systems and boost the overproduction of 
complex natural products.

Copyright © 2022 Ma, Yan, Yang, Yao, Wang, Gao and Cui.

DOI: 10.3389/fbioe.2022.1095464
PMCID: PMC9811125
PMID: 36619381

Conflict of interest statement: The authors declare that the research was 
conducted in the absence of any commercial or financial relationships that could 
be construed as a potential conflict of interest.


5. Metab Eng. 2022 Jan;69:198-208. doi: 10.1016/j.ymben.2021.12.002. Epub 2021
Dec  10.

Overproduction of medicinal ergot alkaloids based on a fungal platform.

Yao Y(1), Wang W(2), Shi W(3), Yan R(1), Zhang J(1), Wei G(2), Liu L(4), Che 
Y(5), An C(6), Gao SS(7).

Author information:
(1)State Key Laboratory of Microbial Resources, Institute of Microbiology, 
Chinese Academy of Sciences, Beijing, 100101, PR China.
(2)State Key Laboratory of Microbial Resources, Institute of Microbiology, 
Chinese Academy of Sciences, Beijing, 100101, PR China; University of Chinese 
Academy of Sciences, Beijing, 100049, PR China.
(3)Microbial Resource and Big Data Center, Institute of Microbiology, Chinese 
Academy of Sciences, Beijing, 100101, PR China.
(4)State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy 
of Sciences, Beijing, 100101, PR China.
(5)Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and 
Peking Union Medical College, Beijing, 100050, PR China.
(6)State Key Laboratory of Microbial Resources, Institute of Microbiology, 
Chinese Academy of Sciences, Beijing, 100101, PR China. Electronic address: 
ancy@im.ac.cn.
(7)State Key Laboratory of Microbial Resources, Institute of Microbiology, 
Chinese Academy of Sciences, Beijing, 100101, PR China. Electronic address: 
gaoss@im.ac.cn.

Privileged ergot alkaloids (EAs) produced by the fungal genus Claviceps are used 
to treat a wide range of diseases. However, their use and research have been 
hampered by the challenging genetic engineering of Claviceps. Here we 
systematically refactored and rationally engineered the EA biosynthetic pathway 
in heterologous host Aspergillus nidulans by using a Fungal-Yeast-Shuttle-Vector 
protocol. The obtained strains allowed the production of diverse EAs and related 
intermediates, including prechanoclavine (PCC, 333.8 mg/L), chanoclavine (CC, 
241.0 mg/L), agroclavine (AC, 78.7 mg/L), and festuclavine (FC, 99.2 mg/L), etc. 
This fungal platform also enabled the access to the methyl-oxidized EAs (MOEAs), 
including elymoclavine (EC), lysergic acid (LA), dihydroelysergol (DHLG), and 
dihydrolysergic acid (DHLA), by overexpressing a P450 enzyme CloA. Furthermore, 
by optimizing the P450 electron transfer (ET) pathway and using multi-copy of 
cloA, the titers of EC and DHLG have been improved by 17.3- and 9.4-fold, 
respectively. Beyond our demonstration of A. nidulans as a robust platform for 
EA overproduction, our study offers a proof of concept for engineering the 
eukaryotic P450s-contained biosynthetic pathways in a filamentous fungal host.

Copyright © 2021. Published by Elsevier Inc.

DOI: 10.1016/j.ymben.2021.12.002
PMID: 34902590 [Indexed for MEDLINE]