New biotechnology最新文献

{"title":"tiRNA: An efficient and controllable gene silencing technology via translation inhibition.","authors":"Bei Xia, Jiajing Cai, Zhilin He, Qubo Zhu","doi":"10.1016/j.nbt.2025.07.010","DOIUrl":"10.1016/j.nbt.2025.07.010","url":null,"abstract":"<p><p>RNA-targeted therapies have emerged as a revolutionary breakthrough in biomedicine recently, offering unprecedented precision in regulating gene expression. Among these, steric blocking oligonucleotides (SBOs) represent a unique class of therapeutics that function through a steric blocking mechanism, allowing for reversible effects without RNA degradation. However, designing SBOs is challenging due to the lack of universal guidelines and the need for detailed analysis of mRNA/pre-mRNA characteristics, key site distributions, and RNA-binding protein interactions. To address this, we developed a novel aptamer-based technology called translation inhibition RNA (tiRNA), which inhibits mRNA translation by linking an eIF4G-targeting aptamer to a reverse complementary sequence of the target gene's 5'-UTR. This approach leverages the typical translation initiation mechanism of mRNA and employs a straightforward design strategy to specifically target and inhibit the translation of selected mRNAs without affecting others or causing RNA degradation. The efficacy of tiRNA is comparable to that of siRNA, providing precision, safety, and controllability for treating diseases linked to protein overexpression. Moreover, the effects of tiRNA can be reversed using a specially designed neutralizing strand, restoring normal mRNA translation and enhancing treatment controllability and personalization. This method holds significant potential for applications in cancer, gene therapy, and other fields requiring precise regulation of protein expression.</p>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":" ","pages":"177-190"},"PeriodicalIF":4.9,"publicationDate":"2025-11-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144799697","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Engineering enhanced signal peptides: A high-throughput computational pipeline for optimizing therapeutic protein production in CHO cells","authors":"Ji-Hong Zhang ,&nbsp;Chong Wang ,&nbsp;Zhao-Yang Wu ,&nbsp;Chun-Liu Mi ,&nbsp;Zi-Meng Han ,&nbsp;Hui-Xian Dong ,&nbsp;Tian-Yun Wang","doi":"10.1016/j.nbt.2025.10.007","DOIUrl":"10.1016/j.nbt.2025.10.007","url":null,"abstract":"<div><div>Rational design of signal peptides (SPs), crucial for efficient therapeutic protein secretion in Chinese hamster ovary (CHO) cells, remains challenging due to their context-dependency activity. To overcome this limitation and enable the discovery of novel high-performance SPs, we developed a high-throughput computational screening pipeline. This approach leverages the deep learning model SignalP 6.0 to screen millions of SP variants derived from diverse mouse/human wild-type libraries and C-region mutants. As a demonstration of its broad applicability, we applied this pipeline to optimize SPs for human serum albumin (HSA) expression. Ranking candidates based on predicted translocation efficiency and cleavage accuracy identified thirty promising SPs outperforming native HSA SP. Experimental validation in CHO cells confirmed multiple novel SPs that significantly enhanced HSA yields, both transiently (e.g., M1_MATN2, 1.93-fold; H5_CXL14, 1.63-fold) and stably (e.g., H5_CXL14, 2.89-fold; M1_MATN2, 1.86-fold). Crucially, our analysis revealed novel insights: hydropathicity profiling uncovered a distinctive and highly effective signature in the top high-performing H5_CXL14 SP, characterized by rapid hydrophobic onset, a continuous highly hydrophobic core, and peak hydrophobicity. Solubility predictions suggested wild-type SPs enhanced secreted protein solubility, while C-region mutants had neutral or negative effects. Furthermore, a novel correlation was observed between high-expression levels and more stable mRNA secondary structures (lower minimum free energy, MFE). This integrated computational-experimental pipeline represents a significant advance, enabling the rational design of protein-specific SP with high efficiency. It drastically reduces the experimental screening burden and holds substantial promise for broadly optimizing therapeutic protein production platforms, as demonstrated here for HSA in CHO cells.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 223-235"},"PeriodicalIF":4.9,"publicationDate":"2025-10-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145337343","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Establishing a cutting-edge protoplast technology platform for applying new genomic techniques in Cichorium spp","authors":"Adriana Fernanda Soria Garcia ,&nbsp;Silvia Farinati ,&nbsp;Samela Draga,&nbsp;Damiano Riommi,&nbsp;Giovanni Gabelli,&nbsp;Alessandro Vannozzi,&nbsp;Gianni Barcaccia,&nbsp;Fabio Palumbo","doi":"10.1016/j.nbt.2025.10.008","DOIUrl":"10.1016/j.nbt.2025.10.008","url":null,"abstract":"<div><div>Genome editing technologies, especially those based on the CRISPR/Cas9 system, have revolutionized crop breeding by enabling precise genetic modifications. Specifically, delivering preassembled ribonucleoprotein (RNP) complexes—consisting of the Cas9 endonuclease coupled to specific single guide RNAs (sgRNAs)—into protoplasts offers an effective DNA-free method that prevents the integration of foreign genetic material. Despite the availability of detailed protocols, establishing a standardized and efficient <em>in vitro</em> regeneration procedure—from protoplast isolation to whole plant regeneration—remains challenging due to significant variability in regeneration efficiency across different varieties and biotypes. Therefore, optimizing each step is essential to maximize the recovery of successful edited plants. In this study, we developed an efficient protocol for regenerating whole plants from protoplasts isolated from 12 representative Italian varieties of chicory and endive. We focused on leaf chicory and endive biotypes with high horticultural value, including Radicchio types, which are important targets for quality improvement. Our optimized platform supports protoplast isolation, PEG-mediated transfection, and plant regeneration, demonstrating promising potential for future genome editing applications. Notably, the high responsiveness of protoplasts to PEG-mediated transfection suggests that coupling this method with our regeneration procedure could facilitate the use of advanced biotechnological strategies. The combination of high transient transformation efficiency, versatile encapsulation techniques, and successful plant regeneration establishes chicory and endive as promising candidates for DNA-free genome editing via protoplasts, providing a technically precise approach with reduced environmental and economic impacts compared to conventional breeding methods.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 206-222"},"PeriodicalIF":4.9,"publicationDate":"2025-10-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145318605","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cultivation at a liquid–liquid interface prevents phenotypic heterogeneity of human induced pluripotent stem-derived retinal pigment epithelial cells","authors":"Rie Sonoi ,&nbsp;Masamichi Kamihira","doi":"10.1016/j.nbt.2025.10.006","DOIUrl":"10.1016/j.nbt.2025.10.006","url":null,"abstract":"<div><div>Culturing retinal pigment epithelial (RPE) cells, a valuable source for drug discovery and cell transplantation therapies, on a polystyrene solid interface often induces spontaneous phenotypic heterogeneity, including cobblestone-shaped, dome-shaped, and stratified cells within a passaged cell population. Understanding and regulating these phenotypic changes is essential for producing high-quality and safe cell sources. In this study, we developed a cultivation strategy to promote the uniform maturation of human induced pluripotent stem (hiPS)-derived RPE cells by focusing on their behavior in a culture vessel. hiPS-RPE cells cultured at the solid–liquid interface exhibited phenotypic heterogeneity, characterized by cobblestone, dome-shaped, and stratified morphologies, indicating RPE phenotype shifts associated with cellular senescence. However, replacing the Rho-associated coiled-coil kinase (ROCK) inhibitor Y27632 with forskolin, which enhances cell-cell and cell-substrate adhesion, facilitated uniform maturation of confluent hiPS-RPE cells on a laminin-332-coated liquid–liquid interface. Quantitative analysis revealed that the levels of tight junction formation, <em>F</em>_Z, and the homogeneity index, i.e., the degree of uniform cell distribution, <em>H</em>_LN, were consistent between the central and peripheral regions of the culture vessel (<em>F</em>_Z = 0.97, <em>H</em>_LN = 0.95). These findings highlight the importance of using a liquid–liquid interface to suppress spontaneous phenotypic heterogeneity by promoting uniform cell distribution. Our study presents a novel methodology for efficiently achieving uniform maturation of functional hiPS-RPE cells at the liquid–liquid interface within a culture vessel.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 196-205"},"PeriodicalIF":4.9,"publicationDate":"2025-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145308701","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Monoclonal antibodies production in microbial systems: Current status, challenges and perspectives","authors":"Di Zhang,&nbsp;Hong Liu,&nbsp;Yaohua Zhong","doi":"10.1016/j.nbt.2025.10.005","DOIUrl":"10.1016/j.nbt.2025.10.005","url":null,"abstract":"<div><div>Monoclonal antibodies (mAbs) serve as indispensable tools in diagnostics, clinical therapeutics, and biomedical research. However, their large-scale production faces significant challenges due to the high costs and lengthy timelines associated with conventional mammalian cell-based expression systems. Microbial expression platforms have emerged as a transformative alternative, offering cost-effectiveness, rapid cultivation cycles, and superior genetic tractability for industrial-scale monoclonal antibodies production. Recent advances in Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9-mediated gene editing enable precise metabolic engineering of host strains to enhance protein folding, secretion efficiency, and translational accuracy. Synthetic biology approaches facilities the reconstruction of mammalian glycosylation pathways in microbial systems, yielding monoclonal antibodies with near-native structural integrity. Furthermore, AI (artificial intelligence)-driven optimization of expression vectors, promoter systems, and culture conditions, combined with high-throughput screening of engineered strains, significantly accelerates the identification of high-yield production clones. This review comprehensively examines current progress in microbial expression systems, strain engineering strategies, and fermentation optimization for enhanced monoclonal antibodies production, while critically discussing existing limitations and potential solutions to advance the field.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 163-173"},"PeriodicalIF":4.9,"publicationDate":"2025-10-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145266027","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comparative transcriptomics of CAZy enzymes in white- and brown-rot agaricomycetes: Evolutionary insights into lignocellulose degradation and the relevance of GH16 glycoside hydrolase functional divergence","authors":"Idoia Jiménez ,&nbsp;Manuel Alfaro ,&nbsp;Edurne Garde ,&nbsp;Gumer Pérez ,&nbsp;Ana Fernandez-Morales ,&nbsp;Anna Lipzen ,&nbsp;Kathleen Lail ,&nbsp;Diane Bauer ,&nbsp;Kerrie Barry ,&nbsp;Igor V. Grigoriev ,&nbsp;Lucía Ramírez ,&nbsp;Antonio G. Pisabarro","doi":"10.1016/j.nbt.2025.10.004","DOIUrl":"10.1016/j.nbt.2025.10.004","url":null,"abstract":"<div><div>White-rot and brown-rot fungi (WRF and BRF, respectively) decompose lignocellulose, the main structural component of plant biomass, through distinct mechanisms. This study examines the transcriptomic responses of three WRF species (<em>Pleurotus ostreatus</em>, <em>Phanerochaete chrysosporium</em>, and <em>Heterobasidion irregulare</em>) and two BRF species (<em>Fomitopsis schrenkii</em> and <em>Rhodonia placenta</em>) grown on poplar wood (W) and glucose (G) as sole carbon sources. RNA-seq analysis revealed upregulation of carbohydrate-active enzymes (CAZymes) linked to lignocellulose degradation. WRF displayed a broader enzymatic repertoire, whereas BRF employed a more selective approach. Among these responses, GH16 glycoside hydrolases were consistently upregulated across all species, including BRF. Since GH16 enzymes are involved in both plant hemicellulose modification and cell wall remodeling, their wood-induced expression may reflect multiple processes rather than a single conserved wood-decay mechanism. Structural and phylogenetic analyses revealed species-specific divergence, consistent with potential functional specialization. These findings broaden our understanding of fungal enzymatic strategies and highlight GH16 enzymes as candidates for further study. Beyond the immediate context of wood decay, this work has broader implications for fungal ecology, evolutionary biology, and biotechnological applications such as biomass conversion and sustainable bioenergy.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 236-256"},"PeriodicalIF":4.9,"publicationDate":"2025-10-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145280736","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A comprehensive approach to optimized ultrasound stimulation for enhanced astaxanthin synthesis in Haematococcus pluvialis: The cultivation strategy","authors":"Ju Yeon Lee ,&nbsp;Jaewon Park ,&nbsp;Sang-Il Han ,&nbsp;Min Seo Jeon ,&nbsp;Yoon-E. Choi","doi":"10.1016/j.nbt.2025.10.001","DOIUrl":"10.1016/j.nbt.2025.10.001","url":null,"abstract":"<div><div>Astaxanthin, a ketocarotenoid, is a potent antioxidant, and the microalga <em>Haematococcus pluvialis</em> is one of the major natural producers of astaxanthin. In this study, a strategy utilizing ultrasound throughout the cultivation process was introduced to induce astaxanthin biosynthesis in <em>H. pluvialis</em>. The effects of various ultrasound intensities, treatment durations, and frequencies on both cell viability and astaxanthin induction were characterized and optimized. The optimized approach was then applied throughout the culture process for each stage, and the physiological effects of long-term ultrasound stimulation on <em>H. pluvialis</em> were analyzed. The results demonstrated a significant increase of 43.2 % in astaxanthin content compared to the control. Ultrasound, as a stress stimulus, not only promoted astaxanthin synthesis but also enhanced its intracellular accumulation by facilitating its distribution throughout the cell. These findings highlight the potential of ultrasound as an effective tool for enhancing astaxanthin production in <em>H. pluvialis</em>, contributing to the sustainable and cost-effective production of high-value antioxidants in the biotechnology industry.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 185-195"},"PeriodicalIF":4.9,"publicationDate":"2025-10-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145275368","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Heterologous production of β-pinene in the chloroplast of the marine diatom Phaeodactylum tricornutum","authors":"Nicola Trevisan ,&nbsp;Michele Fabris ,&nbsp;John van der Oost ,&nbsp;Maria Barbosa ,&nbsp;Sarah D’Adamo","doi":"10.1016/j.nbt.2025.10.003","DOIUrl":"10.1016/j.nbt.2025.10.003","url":null,"abstract":"<div><div>Microalgae are considered promising hosts for the sustainable production of plant-derived secondary metabolites, such as terpenoids. In particular, the model marine diatom <em>Phaeodactylum tricornutum</em> is a promising candidate for the heterologous production of such chemicals due to its robustness, relatively fast growth, and high natural accumulation of terpenoids. In addition, <em>P. tricornutum</em> possesses two separate pathways for the synthesis of terpenoid precursors, the MEP pathway in the chloroplast and the MVA pathway in the cytosol. In this study, we explored the two pathways for the heterologous β-pinene production in <em>P. tricornutum</em> by episomally expressing pinene synthase in either the chloroplast or cytosol for the first time. Our results show that the chloroplast expression from episomes led to β-pinene titers of up to 10.27 ± 1.45 µg<strong>·</strong>L⁻¹. No β-pinene synthesized from the cytosolic MVA pathway precursors was detected, however the expression and functionality of the pinene synthase was confirmed both in the cytosol and in the chloroplast. Furthermore, to enhance production in the chloroplast, we developed more stable transgenic lines with random chromosomal integration of two different pinene synthase genes. We observed higher titers compared to the episomal mutants, up to 19.35 ± 1.42 µg<strong>·</strong>L⁻¹ with the pinene synthase from <em>Abies grandis</em> and 20.07 ± 0.51 µg<strong>·</strong>L⁻¹ with the pinene synthase from <em>Citrus limon</em>. All the β-pinene producing strains used in this study also produced α-pinene as a side product, which accounted for 20–25 % of total monoterpenoid production. Overall, this study represents a fundamental step in microalgal engineering towards the synthesis of monoterpenoids.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 174-184"},"PeriodicalIF":4.9,"publicationDate":"2025-10-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145266026","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Site-specifically immobilized D-amino acid dehydrogenase for the synthesis of D-phenylalanine","authors":"Krisztina Boros ,&nbsp;András-Ernő Iszlai ,&nbsp;Laura-Edit Barabás,&nbsp;László Csaba Bencze,&nbsp;Monica Ioana Toşa","doi":"10.1016/j.nbt.2025.09.006","DOIUrl":"10.1016/j.nbt.2025.09.006","url":null,"abstract":"<div><div>Aromatic <span>d</span>-amino acids (<span>d</span>-AAs) have gained increasing attention as chiral building blocks, with biocatalytic procedures emerging as powerful methods for their asymmetric synthesis. <span>d</span>-Amino acid dehydrogenases (<span>D</span>AADH), developed by protein engineering from <em>meso</em>-diaminopimelate dehydrogenases, step out as highly efficient biocatalysts for the reductive amination-based production of <span>d</span>-AAs. Enzyme immobilization allows the recovery and reuse of biocatalysts, while also enhances their operational stability, which is essential for industrial applications. Since the immobilization of DAADHs have been less explored, we targeted to further progress within the covalent immobilization of DAADH from <em>Ureibacillus thermosphaericus</em>, by exploring its site-specific, covalent immobilization. The individual replacement of several surficial Ser residues to Cys at positions 2, 58, 92, 185, 192 and 317 of <em>Ut</em>DAADH, allowed their site-specific immobilization onto the maleimide-functionalized Purolite® ECR8415F methacrylic support. The highest specific activity values, <span><math><mo>∼</mo></math></span>0.078 and <span><math><mo>∼</mo></math></span>0.083 U/mg provided by immobilization through Cys2 and Cys192, respectively, showed 2.1- and 2.2-fold higher values compared to the covalently, but non-specifically immobilized <em>Ut</em>DAADH. The recyclability of the immobilized preparations was tested among 10 reductive amination-cycles of phenylpyruvate and based on the retained conversion and specific activities, <em>Ut</em>DAADH immobilized through S2C was the best-performing biocatalyst, maintaining 65–70 % conversion and 50 % of the initial activity after the 10^th cycle. After the assessment of optimal enzyme/substrate <em>ratio</em>, the immobilized S2C <em>Ut</em>DAADH was tested in three consecutive 200 mg-scale reaction, providing the enantiopure <span>d</span>-Phe with complete conversions and excellent &gt; 88 % isolation yields, supporting its synthetic applicability.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 145-154"},"PeriodicalIF":4.9,"publicationDate":"2025-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145213212","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Prenylation of diverse indole derivatives by the fungal aromatic prenyltransferase RePT","authors":"Pimvisuth Chunkrua,&nbsp;Mirjam A. Kabel,&nbsp;Jean-Paul Vincken,&nbsp;Willem J.H. van Berkel,&nbsp;Wouter J.C. de Bruijn","doi":"10.1016/j.nbt.2025.09.002","DOIUrl":"10.1016/j.nbt.2025.09.002","url":null,"abstract":"<div><div>Prenylation is a widespread natural modification of compounds that serves to functionalize and often enhance the bioactivity of plant and microbial secondary metabolites, including indole derivatives. In this study, we aimed to expand the library of prenylated indoles using RePT, a fungal (i.e. <em>Rasamsonia emersonii</em>) aromatic prenyltransferase from the dimethylallyl tryptophan synthase (DMATS) family. Previous work showed that RePT readily <em>C</em>7- and <em>N</em>1-prenylated <span>l</span>-tryptophan, and <em>O</em>-prenylated <span>l</span>-tyrosine and a number of phenolic stilbenes. Here, we investigated its regioselectivity further with 23 indole substrates, including tryptophan derivatives with varying <em>C</em>4-<em>C</em>7 substituents and several <em>C</em>3-substituted indoles. High conversion was observed primarily with fluorinated tryptophans and unsubstituted indole. Product analysis by UHPLC-PDA-ESI-MSⁿ and NMR revealed that RePT mainly catalyzed either normal prenylation at <em>C</em>7 or reverse prenylation at <em>N</em>1 on a series of halogenated tryptophans. The regioselectivity observed for several substrates was strongly influenced by the position of the halogen substituent, particularly fluorine, which displayed its characteristic <em>ortho-</em>/<em>para-</em>directing effect. In the absence of the amino acid moiety, RePT’s regioselectivity in some cases shifted from its typical preference, leading to prenylation at alternative positions such as <em>C</em>3 and <em>C</em>6. These findings showcase the versatility of RePT for modifying diverse indole derivatives and demonstrate, for the first time, halogen-induced steering of the regioselectivity of DMATS to facilitate synthesis of bioactive prenylated compounds and intermediates.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"90 ","pages":"Pages 88-96"},"PeriodicalIF":4.9,"publicationDate":"2025-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145086626","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}