Biosensor and machine learning-aided engineering of an amaryllidaceae enzyme

Nature Communications, Год журнала: 2024, Номер 15(1)

Опубликована: Март 7, 2024

Abstract A major challenge to achieving industry-scale biomanufacturing of therapeutic alkaloids is the slow process biocatalyst engineering. Amaryllidaceae alkaloids, such as Alzheimer’s medication galantamine, are complex plant secondary metabolites with recognized value. Due their difficult synthesis they regularly sourced by extraction and purification from low-yielding daffodil Narcissus pseudonarcissus . Here, we propose an efficient biosensor-machine learning technology stack for development, which apply engineer enzyme in Escherichia coli Directed evolution used develop a highly sensitive (EC 50 = 20 μM) specific biosensor key alkaloid branchpoint 4’-O-methylnorbelladine. structure-based residual neural network (MutComputeX) subsequently developed generate activity-enriched variants methyltransferase, rapidly screened biosensor. Functional identified that yield 60% improvement product titer, 2-fold higher catalytic activity, 3-fold lower off-product regioisomer formation. solved crystal structure elucidates mechanism behind beneficial mutations.

Язык: Английский

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Biosynthesis of plant neuroactive alkaloids treating Alzheimer’s disease

Frontiers in Pharmacology, Год журнала: 2025, Номер 16

Опубликована: Фев. 5, 2025

Alzheimer's disease (AD) is a debilitating neurodegenerative disorder characterized by cognitive decline and memory impairment. With increasing global prevalence, the need for effective therapeutic interventions critical. Among currently approved treatments, acetylcholinesterase inhibitors (AChEIs) like huperzine A galantamine stand out due to their neuroprotective roles. These plant-derived alkaloids have demonstrated significant efficacy in alleviating symptoms acetylcholine levels brain.While numerous other plant exhibit varying degrees of neuroactive properties, remain only marketed as specific treatments AD diseases. For example, such berberine (from Berberis species) rhynchophylline Uncaria rhynchophylla) shown potential targeting amyloid-beta (Aβ) aggregation, oxidative stress, tau hyperphosphorylation. Similarly, harmine has ability inhibit hyperphosphorylation through dual inhibition glycogen synthase kinase-3 beta (GSK-3β) specificity tyrosine phosphorylation regulated kinase 1A (DYRK1A). However, these metabolites not yet advanced clinical applications (Ng et al., 2015;Rezaul Islam 2024).The elucidation biosynthetic pathways marks advancement understanding biochemistry specialized metabolism. It advances our but also provides opportunities sustainable scalable production synthetic biology approaches. By leveraging this approach, researchers can reconstruct natural products microbial or systems, facilitating efficient reducing reliance on native sources valuable compounds (Liu 2023;Zhang 2023;Bai 2024a;Teng 2024). This opinion highlights implications discoveries future research application treatment.Huperzine A, derived from Huperzia serrata (Lycopodiaceae), well-known AChEI that been widely used traditional Chinese medicine (Ma Gang, 2004;Yang 2017;Wang 2020;Zhang The pathway provided crucial insights into formation Lycopodium uncovered enzymes with novel functions (Li 2022;Ushimaru Abe, 2023;Cheng Recent studies identified three neofunctionalized α-carbonic anhydrase-like (CAL) responsible key Mannich-like condensations form core carbon-carbon bonds alkaloids, steps construction polycyclic skeletons.Through transcriptome analysis enzyme characterization, Nett al. CAL-1 CAL-2, which promote annulation reactions (Nett 2023;Liu 2024a;Zamar proceeds stereospecific modifications scaffold tailoring, involving additional Fe(II)-dependent dioxygenases, introduce oxidation final bioactive (Figure 1) 2021;Nett 2023;Ushimaru 2023). findings shed light complex evolution species, suggesting evolved metabolite defense mechanism. breakthrough deepens opens door bioengineering chassis, resources (Zhang 2022;Gao 2024b;Golubova 2024).Galantamine, an alkaloid plants Amaryllidaceae family, particularly daffodils (Narcissus spp.), another treatment (Prvulovic 2010). Similar was recently elucidated, providing invaluable its (Kilgore 2014;Li 2018;Li 2019;Hu 2021;Mehta discovery began identifying precursor, 4'-O-methylnorbelladine (4OMN), followed coupling catalyzed cytochrome P450 NtCYP96T6. facilitates para-ortho (p-o') necessary produce skeleton. Subsequent methylation reduction steps, NtNMT1 NtAKR1 respectively, complete biosynthesis 2) (Mehta profound metabolic engineering. sourced primarily populations daffodils, biosynthesize it engineered systems holds promise Additionally, galantamine's helps understand how generate chemical diversity simple precursors, foundation engineering related value.The underscores complexity elegance Both share common feature acting inhibitors, though evolutionary origins differ significantly. families, distinct pressures, developed highly allow synthesize intricate structures. While represents advancement, several challenges remain.First, vivo functional roles are fully understood. speculated they serve against herbivores, regulatory mechanisms governing elusive (Chavez Further ecological could provide important medicinal plants, pathways, interactions environment (Szypuła Pietrosiuk, 2024).Second, scalability producing heterologous remains challenge. transient expression N.benthamiana proof-of-concept biosynthesis, translating industrial-scale will require optimization gene expression, precursor supply, enzymatic activity plant-based platforms 2024b;Yang Optimizing enhancing activity, achieving high-yield critical bottlenecks.Microbial platforms, Saccharomyces cerevisiae Pichia pastoris, offer promising avenues large-scale ease genetic manipulation 2023;Yang On hand, chassis Nicotiana benthamiana unique advantages, including environments compartmentalized cells conducive 2023;Golubova 2024;Liu 2024b). Advances CRISPR-based genome editing, multi-gene assembly, flux pivotal overcoming current limitations (Liao 2023;Xie 2023;Teng tools, create paving way accessible therapeutics disease.Finally, discovering new species should be overlooked. likely represent fraction produce. Systematic exploration yield AChE diseases.The moment neuropharmacology. deepen metabolism practical treatments. As population ages burden diseases grows, plantderived continue play essential role treatment. lies intersection biology, engineering, sciences, innovative solutions neurological disorders.MY: Conceptualization, Writing-original draft. MY, QY, ZZ: Writing-review editing.

Язык: Английский

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Elucidation and biosynthesis of tetrahydroisoquinoline alkaloids: Recent advances and prospects

Biotechnology Advances, Год журнала: 2025, Номер 79, С. 108524 - 108524

Опубликована: Янв. 28, 2025

Язык: Английский

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Elucidating the enzyme network driving Amaryllidaceae alkaloids biosynthesis in Leucojum aestivum

Plant Biotechnology Journal, Год журнала: 2025, Номер unknown

Опубликована: Март 4, 2025

Summary Amaryllidaceae alkaloids (AAs) are diverse bioactive metabolites with significant pharmaceutical potential, derived from 4′‐O‐methylnorbelladine (4′ O M). The biosynthesis of these compounds involves the condensation tyramine and 3,4‐dihydroxybenzaldehyde by norbelladine synthase (NBS) and/or noroxomaritidine/norcraugsodine reductase (NR), followed ‐methylation. Cytochrome P450 enzymes, particularly CYP96T family, introduce further structural diversity through C–C couplings, resulting in lycorine, galanthamine crinine cores. Despite their importance, exact biosynthetic pathways remain poorly defined. In this study, we describe key enzymes Leucojum aestivum ( La ), providing crucial insight into AA biosynthesis. Transient expression Nicotiana benthamiana demonstrated that NBS NRII catalyse conversion to norbelladine, which is subsequently ‐methylated a norbelladine‐4′‐ ‐methyltransferase N4′ MT) planta . Co‐agroinfiltration NBS, NRII, MT CYP96T1 resulted production various phenol‐coupled products, lycorine as predominant compound, alongside haemanthamine, crinine/vittatine norgalanthamine. This study identifies CYP96T2 first monocot capable catalysing all three regioselective C‐C phenol couplings also highlights substrate promiscuity NRII. findings not only elucidate critical steps but open new avenues for biotechnological application producing valuable alkaloids, offering potential novel drug development.

Язык: Английский

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Coclaurine N-Methyltransferase-Like Enzymes Catalyze the Last Reaction of the Biosynthesis of the Anti-Alzheimer's Drug Galanthamine in Amaryllidaceae

Опубликована: Янв. 1, 2025

Язык: Английский

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A concise enzyme cascade enables the manufacture of natural and halogenated protoberberine alkaloids

Nature Communications, Год журнала: 2025, Номер 16(1)

Опубликована: Фев. 23, 2025

Язык: Английский

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Toward an integrated omics approach for plant biosynthetic pathway discovery in the age of AI

Trends in Biochemical Sciences, Год журнала: 2025, Номер unknown

Опубликована: Фев. 1, 2025

Язык: Английский

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Synthetic Biology in Natural Product Biosynthesis

Chemical Reviews, Год журнала: 2025, Номер unknown

Опубликована: Март 21, 2025

Synthetic biology has played an important role in the renaissance of natural products research during post-genomics era. The development and integration new tools have transformed workflow product discovery engineering, generating multidisciplinary interest field. In this review, we summarize recent developments biosynthesis from three different aspects. First, advances bioinformatics, experimental, analytical to identify associated with predicted biosynthetic gene clusters (BGCs) will be covered. This followed by extensive review on heterologous expression bacterial, fungal plant organisms. native host-independent paradigm identification, pathway characterization, enzyme is where synthetic most prominent role. Lastly, strategies engineer pathways for structural diversification complexity generation discussed, including assembly-line megasynthase precursor-directed modification, combinatorial biosynthesis.

Язык: Английский

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O-methylation Modifications in the Biosynthetic Pathway of Dibenzocyclooctadiene Lignans

Plant Physiology and Biochemistry, Год журнала: 2025, Номер unknown, С. 109863 - 109863

Опубликована: Апрель 1, 2025

Язык: Английский

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Artificial intelligence in the discovery and modification of biological elements in medicinal plants

Medicinal Plant Biology, Год журнала: 2025, Номер 4(1), С. 0 - 0

Опубликована: Янв. 1, 2025

Язык: Английский

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