RESUMEN
This study investigated the potential of endophytic fungi to produce paclitaxel (Taxol®), a potent anticancer compound widely employed in chemotherapy. This research aimed to identify, confirm, and characterize endophytic fungi capable of paclitaxel (PTX) production and assess their paclitaxel yield. Additionally, it aimed to investigate factors influencing paclitaxel production. A total of 100 endophytic fungal isolates were collected and identified from the roots of Artemisia judaica. Aspergillus fumigatiaffinis exhibited the highest PTX production (26.373 µg L-1) among the isolated endophytic fungi. The strain was identified as A. fumigatiaffinis (Accession No. PP235788.1). Molecular identification confirmed its novelty, representing the first report of PTX production by A. fumigatiaffinis, an endophyte of Artemisia judaica. Optimization through full factorial design of experiments (DOE) and response surface methodology (RSM) significantly enhanced PTX production to 110.23 µg L-1 from 1 g of dry weight of the fungal culture under optimal conditions of pH 8.0, 150 µg L-1 becozyme supplementation, and 18 days of fermentation in potato dextrose broth. The presence of paclitaxel was confirmed using thin layer chromatography, high performance liquid chromatography, and gas chromatography-mass spectrometry. These findings maximize the role of endophytic fungus to produce a secondary metabolite that might be able to replace the chemically produced PTX and gives an opportunity to provide a sustainable source of PTX eco-friendly at high concentrations. KEY POINTS: ⢠Endophytic fungi, like A. fumigatiaffinis, show promise for eco-friendly paclitaxel production ⢠Optimization strategies boost paclitaxel yield significantly, reaching 110.23 µg L -1 ⢠Molecular identification confirms novelty, offering a sustainable PTX source.
Asunto(s)
Aspergillus , Endófitos , Fermentación , Paclitaxel , Paclitaxel/biosíntesis , Aspergillus/metabolismo , Aspergillus/genética , Endófitos/metabolismo , Endófitos/genética , Endófitos/aislamiento & purificación , Endófitos/clasificación , Raíces de Plantas/microbiología , Medios de Cultivo/química , Cromatografía de Gases y Espectrometría de Masas , Cromatografía Líquida de Alta PresiónRESUMEN
Alternaria alternata fungus is a potent paclitaxel producer isolated from Corylus avellana. The major challenge is the lack of optimized media for endophytic fungi productivity. In the effort to maximize the production of taxoids by A. alternata, several fermentation conditions, including pH (pH 4.0-7.0), different types and concentrations of carbon (fructose, glucose, sucrose, mannitol, sorbitol, and malt extract), and nitrogen (urea, ammonium nitrate, potassium nitrate, ammonium phosphate, and ammonium sulfate) were applied step by step. Based on the results, A. alternata in a medium containing sucrose 5% (w/v) and ammonium phosphate 2.5 mM at pH 6.0 showed a rapid and sustainable growth rate, the highest paclitaxel yield (94.8 µg gFW-1 vs 2.8 µg gFW-1 in controls), and the maximum content of amino acids. Additionally, the effect of pectin was evaluated on fungus, and mycelia harvested. Pectin significantly enhanced the growth and taxoid yield on day 21 (respectively 171% and 116% of their corresponding on day 7). The results were checked out by mathematical modeling as well. Accordingly, these findings suggest a low-cost, eco-friendly, and easy-to-produce approach with excellent biotechnological potential for the industrial manufacture of taxoids.
Asunto(s)
Alternaria , Medios de Cultivo , Fermentación , Paclitaxel , Pectinas , Alternaria/metabolismo , Pectinas/metabolismo , Medios de Cultivo/química , Paclitaxel/biosíntesis , Paclitaxel/metabolismo , Modelos Teóricos , Concentración de Iones de Hidrógeno , Nitrógeno/metabolismoRESUMEN
Paclitaxel, a rare diterpene extracted from the bark of Chinese yew (Taxus chinensis), is renowned for its anti-cancer activity and serves as a primary drug for treating cancers. Due to the exceptionally low content of paclitaxel in the bark, a semi-synthetic method that depletes Chinese yew resources is used in the production of paclitaxel, which, however, fails to meet the escalating clinical demand. In recent years, researchers have achieved significant progress in heterologous biosynthesis and metabolic engineering for the production of paclitaxel. This article comprehensively reviews the advancements in paclitaxel production, encompassing chemical synthesis, heterologous biosynthesis, and cell engineering. It provides an in-depth introduction to the biosynthetic pathway and transcriptional regulation mechanisms of paclitaxel, aiming to provide a valuable reference for further research on paclitaxel biosynthesis.
Asunto(s)
Paclitaxel , Paclitaxel/biosíntesis , Ingeniería Metabólica/métodos , Taxus/genética , Taxus/metabolismo , Antineoplásicos Fitogénicos/biosíntesis , Antineoplásicos Fitogénicos/farmacología , Transcripción Genética , Vías Biosintéticas/genéticaRESUMEN
Oxetane synthase (TmCYP1), a novel cytochrome P450 enzyme from Taxus×media cell cultures, has been functionally characterized to efficiently catalyse the formation of the oxetane ring in tetracyclic taxoids. Transient expression of TmCYP1 in Nicotiana benthamiana using 2α,5α,7ß,9α,10ß,13α-hexaacetoxytaxa-4(20),11(12)-diene (1) as a substrate led to the production of a major oxetane derivative, 1ß-dehydroxybaccatin IV (1 a), and a minor 4ß,20-epoxide derivative, baccatin I (1 b). However, feeding the substrate decinnamoyltaxinine J (2), a 5-deacetylated derivative of 1, yielded only 5α-deacetylbaccatin I (2 b), a 4ß,20-epoxide. A possible reaction mechanism was proposed on the basis of substrate-feeding, 2H and 18O isotope labelling experiments, and density functional theory calculations. This reaction could be an intramolecular oxidation-acetoxyl rearrangement and the construction of the oxetane ring may occur through a concerted process; however, the 4ß,20-epoxide might be a shunt product. In this process, the C5-O-acetyl group in substrate is crucial for the oxetane ring formation but not for the 4(20)-epoxy ring formation by TmCYP1. These findings provide a better understanding of the enzymatic formation of the oxetane ring in paclitaxel biosynthesis.
Asunto(s)
Sistema Enzimático del Citocromo P-450 , Éteres Cíclicos , Paclitaxel , Sistema Enzimático del Citocromo P-450/metabolismo , Paclitaxel/biosíntesis , Paclitaxel/química , Paclitaxel/metabolismo , Éteres Cíclicos/química , Éteres Cíclicos/metabolismo , Taxus/enzimología , Taxus/metabolismo , Biocatálisis , Nicotiana/metabolismo , Nicotiana/enzimología , Estructura MolecularRESUMEN
Incomplete understanding of the biosynthetic pathway of the anticancer compound Taxol hinders its production by metabolic engineering. Recent reports by Jiang et al. and other groups now describe the missing steps in Taxol biosynthesis, notably including oxetane ring formation. These findings will promote the sustainable production of Taxol through synthetic biology.
Asunto(s)
Ingeniería Metabólica , Paclitaxel , Biología Sintética , Paclitaxel/biosíntesis , Paclitaxel/metabolismo , Biología Sintética/métodos , Ingeniería Metabólica/métodos , Vías BiosintéticasRESUMEN
Paclitaxel is a well known anticancer compound. Its biosynthesis involves the formation of a highly functionalized diterpenoid core skeleton (baccatin III) and the subsequent assembly of a phenylisoserinoyl side chain. Despite intensive investigation for half a century, the complete biosynthetic pathway of baccatin III remains unknown. In this work, we identified a bifunctional cytochrome P450 enzyme [taxane oxetanase 1 (TOT1)] in Taxus mairei that catalyzes an oxidative rearrangement in paclitaxel oxetane formation, which represents a previously unknown enzyme mechanism for oxetane ring formation. We created a screening strategy based on the taxusin biosynthesis pathway and uncovered the enzyme responsible for the taxane oxidation of the C9 position (T9αH1). Finally, we artificially reconstituted a biosynthetic pathway for the production of baccatin III in tobacco.
Asunto(s)
Alcaloides , Sistema Enzimático del Citocromo P-450 , Ingeniería Metabólica , Paclitaxel , Proteínas de Plantas , Taxoides , Taxus , Alcaloides/biosíntesis , Alcaloides/genética , Hidrocarburos Aromáticos con Puentes/química , Hidrocarburos Aromáticos con Puentes/metabolismo , Éteres Cíclicos/química , Éteres Cíclicos/metabolismo , Paclitaxel/biosíntesis , Taxoides/metabolismo , Taxus/enzimología , Taxus/genética , Sistema Enzimático del Citocromo P-450/química , Sistema Enzimático del Citocromo P-450/genética , Proteínas de Plantas/química , Proteínas de Plantas/genéticaRESUMEN
BACKGROUND: Taxol from Taxus species is a precious drug used for the treatment of cancer and can effectively inhibit the proliferation of cancer cells. However, the growth of Taxus plants is very slow and the content of taxol is quite low. Therefore, it is of great significance to improve the yield of taxol by modern biotechnology without destroying the wild forest resources. Endophytic fungus which symbiosis with their host plants can promote the growth and secondary metabolism of medicinal plants. RESULTS: Here, an endophytic fungus KL27 was isolated from T. chinensis, and identified as Pseudodidymocyrtis lobariellae. The fermentation broth of KL27 (KL27-FB) could significantly promote the accumulation of taxol in needles of T. chinensis, reaching 0.361 ± 0.082 mg/g·DW (dry weight) at 7 days after KL27-FB treatment, which is 3.26-fold increase as compared to the control. The RNA-seq and qRT-PCR showed that KL27-FB could significantly increase the expression of key genes involved in the upstream pathway of terpene synthesis (such as DXS and DXR) and those in the taxol biosynthesis pathway (such as GGPPS, TS, T5OH, TAT, T10OH, T14OH, T2OH, TBT, DBAT and PAM), especially at the early stage of the stimulation. Moreover, the activation of jasmonic acid (JA) biosynthesis and JA signal transduction, and its crosstalk with other hormones, such as gibberellin acid (GA), ethylene (ET) and salicylic acid (SA), explained the elevation of most of the differential expressed genes related to taxol biosynthesis pathway. Moreover, TF (transcriptional factor)-encoding genes, including MYBs, ethylene-responsive transcription factors (ERFs) and basic/helix-loop-helix (bHLH), were detected as differential expressed genes after KL27-FB treatment, further suggested that the regulation of hormone signaling on genes of taxol biosynthesis was mediated by TFs. CONCLUSIONS: Our results indicated that fermentation broth of endophytic fungus KL27-FB could effectively enhance the accumulation of taxol in T. chinensis needles by regulating the phytohormone metabolism and signal transduction and further up-regulating the expression of multiple key genes involved in taxol biosynthesis. This study provides new insight into the regulatory mechanism of how endophytic fungus promotes the production and accumulation of taxol in Taxus sp.
Asunto(s)
Ascomicetos/fisiología , Endófitos/fisiología , Regulación de la Expresión Génica de las Plantas , Paclitaxel/biosíntesis , Reguladores del Crecimiento de las Plantas/metabolismo , Transducción de Señal , Taxus/metabolismo , Genes de Plantas , Paclitaxel/metabolismo , Taxus/microbiología , Regulación hacia ArribaRESUMEN
Taxol, a natural product derived from Taxus, is one of the most effective natural anticancer drugs and the biosynthetic pathway of Taxol is the basis of heterologous bio-production. Here, we report a high-quality genome assembly and annotation of Taxus yunnanensis based on 10.7 Gb sequences assembled into 12 chromosomes with contig N50 and scaffold N50 of 2.89 Mb and 966.80 Mb, respectively. Phylogenomic analyses show that T. yunnanensis is most closely related to Sequoiadendron giganteum among the sampled taxa, with an estimated divergence time of 133.4-213.0 MYA. As with most gymnosperms, and unlike most angiosperms, there is no evidence of a recent whole-genome duplication in T. yunnanensis. Repetitive sequences, especially long terminal repeat retrotransposons, are prevalent in the T. yunnanensis genome, contributing to its large genome size. We further integrated genomic and transcriptomic data to unveil clusters of genes involved in Taxol synthesis, located on the chromosome 12, while gene families encoding hydroxylase in the Taxol pathway exhibited significant expansion. Our study contributes to the further elucidation of gymnosperm relationships and the Taxol biosynthetic pathway.
Asunto(s)
Cycadopsida/clasificación , Evolución Molecular , Genoma de Planta , Paclitaxel/biosíntesis , Filogenia , Taxus/genéticaRESUMEN
The ancient gymnosperm genus Taxus is the exclusive source of the anticancer drug paclitaxel, yet no reference genome sequences are available for comprehensively elucidating the paclitaxel biosynthesis pathway. We have completed a chromosome-level genome of Taxus chinensis var. mairei with a total length of 10.23 gigabases. Taxus shared an ancestral whole-genome duplication with the coniferophyte lineage and underwent distinct transposon evolution. We discovered a unique physical and functional grouping of CYP725As (cytochrome P450) in the Taxus genome for paclitaxel biosynthesis. We also identified a gene cluster for taxadiene biosynthesis, which was formed mainly by gene duplications. This study will facilitate the elucidation of paclitaxel biosynthesis and unleash the biotechnological potential of Taxus.
Asunto(s)
Antineoplásicos/metabolismo , Vías Biosintéticas/genética , Genoma de Planta , Paclitaxel/biosíntesis , Análisis de Secuencia , Taxus/genética , Taxus/metabolismo , Evolución Molecular , Plantas Medicinales/genética , Plantas Medicinales/metabolismoRESUMEN
Taxol is one of the most effective anticancer drugs in the world that is widely used in the treatments of breast, lung and ovarian cancer. The elucidation of the taxol biosynthetic pathway is the key to solve the problem of taxol supply. So far, the taxol biosynthetic pathway has been reported to require an estimated 20 steps of enzymatic reactions, and sixteen enzymes involved in the taxol pathway have been well characterized, including a novel taxane-10ß-hydroxylase (T10ßOH) and a newly putative ß-phenylalanyl-CoA ligase (PCL). Moreover, the source and formation of the taxane core and the details of the downstream synthetic pathway have been basically depicted, while the modification of the core taxane skeleton has not been fully reported, mainly concerning the developments from diol intermediates to 2-debenzoyltaxane. The acylation reaction mediated by specialized Taxus BAHD family acyltransferases (ACTs) is recognized as one of the most important steps in the modification of core taxane skeleton that contribute to the increase of taxol yield. Recently, the influence of acylation on the functional and structural diversity of taxanes has also been continuously revealed. This review summarizes the latest research advances of the taxol biosynthetic pathway and systematically discusses the acylation reactions supported by Taxus ACTs. The underlying mechanism could improve the understanding of taxol biosynthesis, and provide a theoretical basis for the mass production of taxol.
Asunto(s)
Aciltransferasas/metabolismo , Antineoplásicos/metabolismo , Paclitaxel/biosíntesis , Extractos Vegetales/biosíntesis , Taxus/química , Taxus/enzimología , Acilación , Aciltransferasas/genética , Secuencia de Aminoácidos , Vías Biosintéticas , Hidrocarburos Aromáticos con Puentes/metabolismo , Ligasas/metabolismo , Oxigenasas de Función Mixta/metabolismo , Taxoides/metabolismo , Taxus/clasificación , Taxus/genética , TranscriptomaRESUMEN
Taxus, commonly known as yew, is a well-known gymnosperm with great ornamental and medicinal value. In this study, by assembling a chromosome-level genome of the Himalayan yew (Taxus wallichiana) with 10.9 Gb in 12 chromosomes, we revealed that tandem duplication acts as the driving force of gene family evolution in the yew genome, resulting in the main genes for paclitaxel biosynthesis, i.e. those encoding the taxadiene synthase, P450s, and transferases, being clustered on the same chromosome. The tandem duplication may also provide genetic resources for the nature to sculpt the core structure of taxoids at different positions and subsequently establish the complex pathway of paclitaxel by neofunctionalization. Furthermore, we confirmed that there are two genes in the cluster encoding isoenzymes of a known enzyme in the paclitaxel biosynthetic pathway. The reference genome of the Himalayan yew will serve as a platform for decoding the complete biosynthetic pathway of paclitaxel and understanding the chemodiversity of taxoids in gymnosperms.
Asunto(s)
Genoma de Planta , Paclitaxel/biosíntesis , Taxus/genética , Cromosomas de las Plantas , Evolución Molecular , Duplicación de Gen , Taxoides , Taxus/metabolismo , Secuenciación Completa del GenomaRESUMEN
BACKGROUND: LXYL-P1-2 is the first reported glycoside hydrolase that can catalyze the transformation of 7-b-xylosyl-10-deacetyltaxol (XDT) to 10-deacetyltaxol (DT) by removing the D-xylosyl group at the C7 position. Successful synthesis of paclitaxel by one-pot method combining the LXYL-P1-2 and 10- deacetylbaccatin III-10-b-O-acetyltransferase (DBAT) using XDT as a precursor, making LXYL-P1-2 a highly promising enzyme for the industrial production of paclitaxel. The aim of this study was to investigate the catalytic potential of LXYL-P1-2 stabilized on magnetic nanoparticles, the surface of which was modified by Ni2+-immobilized cross-linked Fe3O4@Histidine. RESULTS: The diameter of matrix was 2040 nm. The Km value of the immobilized LXYL-P1-2 catalyzing XDT (0.145 mM) was lower than that of the free enzyme (0.452 mM), and the kcat/Km value of immobilized enzyme (12.952 mM s 1 ) was higher than the free form (8.622 mM s 1 ). The immobilized form maintained 50% of its original activity after 15 cycles of reuse. In addition, the stability of immobilized LXYL-P1-2, maintained 84.67% of its initial activity, improved in comparison with free form after 30 d storage at 4 C. CONCLUSIONS: This investigation not only provides an effective procedure for biocatalytic production of DT, but also gives an insight into the application of magnetic material immobilization technology.
Asunto(s)
Paclitaxel/biosíntesis , Glicósido Hidrolasas/metabolismo , Cinética , Enzimas Inmovilizadas , Nanopartículas , ImanesRESUMEN
BACKGROUND: Taxol is an efficient anticancer drug accumulated in Taxus species. Pseudotaxus chienii is an important member of Taxaceae, however, the level of six taxoids in P. chienii is largely unknown. RESULTS: High accumulation of 10-DAB, taxol, and 7-E-PTX suggested that P. chienii is a good taxol-yielding species for large-scale cultivation. By the omics approaches, a total of 3,387 metabolites and 61,146 unigenes were detected and annotated. Compared with a representative Taxus tree (Taxus yunnanensis), most of the differentially accumulated metabolites and differential expressed genes were assigned into 10 primary and secondary metabolism pathways. Comparative analyses revealed the variations in the precursors and intermediate products of taxol biosynthesis between P. chienii and T. yunnanensis. Taxusin-like metabolites highly accumulated in P. chienii, suggesting a wider value of P. chienii in pharmaceutical industry. CONCLUSIONS: In our study, the occurrence of taxoids in P. chienii was determined. The differential expression of key genes involved in the taxol biosynthesis pathway is the major cause of the differential accumulation of taxoids. Moreover, identification of a number of differentially expressed transcription factors provided more candidate regulators of taxol biosynthesis. Our study may help to reveal the differences between Pseudotaxus and Taxus trees, and promote resource utilization of the endangered and rarely studied P. chienii.
Asunto(s)
Vías Biosintéticas , Metaboloma , Metabolómica , Paclitaxel/biosíntesis , Plantas Medicinales/metabolismo , Especificidad de la Especie , Taxaceae/metabolismo , Especies en Peligro de Extinción , Variación GenéticaRESUMEN
BACKGROUND: Taxus is a valuable woody species with important medicinal value. The bark of Taxus can produce taxol, a natural antineoplastic drug that is widely used in the treatment of breast, ovarian and lung cancers. However, the low content of taxol in the bark of Taxus can not meet the growing clinical demands, so the current research aims at finding ways to increase taxol production. OBJECTIVE: In this review, the research progress of taxol including the factors affecting the taxol content, biosynthesis pathway of taxol, production of taxol in vitro and the application of multi-omics approaches in Taxus as well as future research prospects will be discussed. RESULTS: The taxol content is not only dependent on the species, age and tissues but is also affected by light, moisture levels, temperature, soil fertility and microbes. Most of the enzymes in the taxol biosynthesis pathway have been identified and characterized. Total chemical synthesis, semi-synthesis, plant cell culture and biosynthesis in endophytic fungi have been explored to product taxol. Multi-omics have been used to study Taxus and taxol. CONCLUSION: Further efforts in the identification of unknown enzymes in the taxol biosynthesis pathway, establishment of the genetic transformation system in Taxus and the regulatory mechanism of taxol biosynthesis and Taxus cell growth will play a significant role in improving the yield of taxol in Taxus cells and plants.
Asunto(s)
Antineoplásicos Fitogénicos/biosíntesis , Investigación Biomédica/tendencias , Paclitaxel/biosíntesis , Taxus , Animales , Antineoplásicos Fitogénicos/química , Antineoplásicos Fitogénicos/uso terapéutico , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Paclitaxel/química , Paclitaxel/uso terapéutico , Taxus/microbiologíaRESUMEN
Taxadien-5α-hydroxylase and taxadien-5α-ol O-acetyltransferase catalyze the oxidation of taxadiene to taxadien-5α-ol and subsequent acetylation to taxadien-5α-yl-acetate in the biosynthesis of the blockbuster anticancer drug, paclitaxel (Taxol®). Despite decades of research, the promiscuous and multispecific CYP725A4 enzyme remains a major bottleneck in microbial biosynthetic pathway development. In this study, an interdisciplinary approach was applied for the construction and optimization of the early pathway in Saccharomyces cerevisiae, across a range of bioreactor scales. High-throughput microscale optimization enhanced total oxygenated taxane titer to 39.0 ± 5.7 mg/L and total taxane product titers were comparable at micro and minibioreactor scale at 95.4 ± 18.0 and 98.9 mg/L, respectively. The introduction of pH control successfully mitigated a reduction of oxygenated taxane production, enhancing the potential taxadien-5α-ol isomer titer to 19.2 mg/L, comparable with the 23.8 ± 3.7 mg/L achieved at microscale. A combination of bioprocess optimization and increased gas chromatography-mass spectrometry resolution at 1 L bioreactor scale facilitated taxadien-5α-yl-acetate detection with a final titer of 3.7 mg/L. Total oxygenated taxane titers were improved 2.7-fold at this scale to 78 mg/L, the highest reported titer in yeast. Critical parameters affecting the productivity of the engineered strain were identified across a range of scales, providing a foundation for the development of robust integrated bioprocess control systems.
Asunto(s)
Hidrocarburos Aromáticos con Puentes/metabolismo , Ingeniería Metabólica , Saccharomyces cerevisiae , Taxoides/metabolismo , Paclitaxel/biosíntesis , Saccharomyces cerevisiae/enzimología , Saccharomyces cerevisiae/genéticaRESUMEN
Momilactones from rice have allelopathic activity, the ability to inhibit growth of competing plants. Transferring momilactone production to other crops is a potential approach to combat weeds, yet a complete momilactone biosynthetic pathway remains elusive. Here, we address this challenge through rapid gene screening in Nicotiana benthamiana, a heterologous plant host. This required us to solve a central problem: diminishing intermediate and product yields remain a bottleneck for multistep diterpene pathways. We increased intermediate and product titers by rerouting diterpene biosynthesis from the chloroplast to the cytosolic, high-flux mevalonate pathway. This enabled the discovery and reconstitution of a complete route to momilactones (>10-fold yield improvement in production versus rice). Pure momilactone B isolated from N. benthamiana inhibited germination and root growth in Arabidopsis thaliana, validating allelopathic activity. We demonstrated the broad utility of this approach by applying it to forskolin, a Hedgehog inhibitor, and taxadiene, an intermediate in taxol biosynthesis (~10-fold improvement in production versus chloroplast expression).
Asunto(s)
Diterpenos/metabolismo , Lactonas/metabolismo , Plantas/metabolismo , Transducción de Señal/genética , Alquenos/farmacología , Arabidopsis/efectos de los fármacos , Arabidopsis/crecimiento & desarrollo , Cloroplastos/genética , Colforsina/farmacología , Citosol/metabolismo , Diterpenos/farmacología , Ácido Mevalónico/metabolismo , Oryza/genética , Paclitaxel/biosíntesis , Hojas de la Planta/química , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/crecimiento & desarrollo , Nicotiana/metabolismoRESUMEN
Taxol (paclitaxel), a plant-derived anticancer drug, has been among the most successful anticancer drugs of natural origin. Endophytic fungi have been proposed as a prominent alternative source for Taxol and its intermediate Baccatin III, however the very low yields remain a hinderance to their commercial utilization. Significant research efforts towards this end are underway globally. Here, we report the results on our earlier reported Taxol-producing endophytic fungus, Fusarium solani from the standpoint of spores as seed inoculum and media selection for enhanced Taxol and baccatin III yields. Spores produced on M1D medium with 94.76% viability were used for further media optimization for Taxol and Baccatin III production in five different liquid media under static and shaker condition at different cultivation days. Taxol and Baccatin III when quantified through competitive inhibition enzyme immunoassay (CIEIA), showed maximum production at 136.3 µg L-1 and 128.3 µg L-1, respectively in the modified flask basal broth (MFBB) under shaking condition. Further, two important genes of this pathway, namely taxane 13α-hydroxylase (T13αH) and 10-deacetylbaccatin III-10-ß-O-acetyltransferase (DBAT) have been identified in this fungus. These findings are hoped to assist in further manipulation and metabolic engineering of the parent F. solani strain towards the enhanced production of Taxol and baccatin III.
Asunto(s)
Acetiltransferasas/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Fusarium/genética , Paclitaxel/biosíntesis , Esporas Fúngicas/genética , Taxoides/metabolismo , Cromatografía Líquida de Alta Presión , Cromatografía Liquida , Medios de Cultivo , Fusarium/enzimología , Fusarium/metabolismo , Espectrometría de Masa por Ionización de Electrospray , Esporas Fúngicas/crecimiento & desarrollo , TemperaturaRESUMEN
Paclitaxel, a tetracyclic diterpenoid compounds, was firstly isolated from the bark of the Pacific yew trees. Currently, as a low toxicity, high efficiency, and broad-spectrum natural anti-cancer drug, paclitaxel has been widely used against ovarian cancer, breast cancer, uterine cancer, and other cancers. As the matter of fact, natural paclitaxel from Taxus species has been proved to be environmentally unsustainable and economically unfeasible. For this reason, researchers from all over the world are devoted to searching for new ways of obtaining paclitaxel. At present, other methods, including artificial cultivation of Taxus plants, microbial fermentation, chemical synthesis, tissue and cell culture have been sought and developed subsequently. Meanwhile, the biosynthesis of paclitaxel is also an extremely attractive method. Unlike other anti-cancer drugs, paclitaxel has its unique anti-cancer mechanisms. Here, the source, production, and anti-cancer mechanisms of paclitaxel were summarized and reviewed, which can provide theoretical basis and reference for further research on the production, anti-cancer mechanisms and utilization of paclitaxel.