Synthesis Of Terpenoids Research Articles

Cloning and characterization of DoMYC2 from Dendrobium officinale

MYC proteins, a group of transcription factors belonging to bHLH super family, play important roles in regulation of metabolism and plant growth. In this study, a novel MYC gene, DoMYC2 (GeneBank accession KJ649471), was isolated from Dendrobium officinale. The full length of DoMYC2 cDNA contains 2351 bp with an open reading frame of 1971 bp encoding 656 amino acids. BlastP analyses showed that its amino acid sequence was homologous with typical MYC proteins in other plant species. Furthermore, expression of DoMYC2 could be induced by MeJA and chitosan, respectively. Overexpression of DoMYC2 led to a decrease in transcript levels of AtHMGR2, AtFPS1 and AtFPS2 in Arabidopsis. Moreover, DoMYC2 was able to bind to the E-box cis-elements in yeast. These results suggest that DoMYC2 participate in terpenoid indole alkaloid synthesis through regulating the expression of key enzymes in MVA pathway.

IncreasedHelicoverpa zea(Boddie) Larval Feeding on Cotton Plants with RNAi Construct CYP82D109 that Blocks Gossypol-related Terpenoid Synthesis

Increased<i>Helicoverpa zea</i>(Boddie) Larval Feeding on Cotton Plants with RNAi Construct CYP82D109 that Blocks Gossypol-related Terpenoid Synthesis

Bioinspired total syntheses of terpenoids.

Nature's highly efficient routes for constructing natural products have inspired chemists to mimic these processes in a laboratory setting. This Perspective presents some recent examples of conceptually different bioinspired total syntheses of complex terpenoids and thereby aims to highlight the vast benefits offered by bioinspired strategies.

ChemInform Abstract: Recent Application of Oxa‐Michael Reaction in Complex Natural Product Synthesis

AbstractReview: [87 refs.

Functional characterization of thiolase-encoding genes from Xanthophyllomyces dendrorhous and their effects on carotenoid synthesis.

BackgroundThe basidiomycetous yeast Xanthophyllomyces dendrorhous has been described as a potential biofactory for terpenoid-derived compounds due to its ability to synthesize astaxanthin. Functional knowledge of the genes involved in terpenoid synthesis would create opportunities to enhance carotenoid production. A thiolase enzyme catalyzes the first step in terpenoid synthesis.ResultsTwo potential thiolase-encoding genes were found in the yeast genome; bioinformatically, one was identified as an acetyl-CoA C-acetyltransferase (ERG10), and the other was identified as a 3-ketoacyl Co-A thiolase (POT1). Heterologous complementation assays in Saccharomyces cerevisiae showed that the ERG10 gene from X. dendrorhous could complement the lack of the endogenous ERG10 gene in S. cerevisiae, thereby allowing cellular growth and sterol synthesis. X. dendrorhous heterozygous mutants for each gene were created, and a homozygous POT1 mutant was also obtained. This mutant exhibited changes in pigment composition and higher ERG10 transcript levels than the wild type strain.ConclusionsThe results support the notion that the ERG10 gene in X. dendrorhous is a functional acetyl-CoA C-acetyltransferase essential for the synthesis of mevalonate in yeast. The POT1 gene would encode a functional 3-ketoacyl Co-A thiolase that is non-essential for cell growth, but its mutation indirectly affects pigment production.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-016-0893-2) contains supplementary material, which is available to authorized users.

Recent advances in terpenoid syntheses from China

Total syntheses of terpenoids play an essential role in the development of synthetic organic chemistry and medicinal science. Chinese synthetic community has made more and more profound contribution to this field and has gradually grown into a major force in the world over the past years. In this review, we shall celebrate the achievement by surveying the recent terpenoid syntheses from China since 2010.

Mono- and sesquiterpene release from tomato (Solanum lycopersicum) leaves upon mild and severe heat stress and through recovery: From gene expression to emission responses

Plants frequently experience heat ramps of various severities, but how and to what degree plant metabolic activity recovers from mild and severe heat stress is poorly understood. In this study, we exposed the constitutive terpene emitter, Solanum. lycopersicum leaves to mild (37 and 41°C), moderate (46°C) and severe (49°C) heat ramps of 5min and monitored foliage photosynthetic activity, lipoxygenase pathway volatile (LOX), and mono- and sesquiterpene emissions and expression of two terpene synthase genes, β-phellandrene synthase and (E)-β-caryophyllene/α-humulene synthase, through a 24h recovery period upon return to pre-stress conditions. Leaf monoterpene emissions were dominated by β-phellandrene and sesquiterpene emissions by (E)-β-caryophyllene, and thus, these two terpene synthase genes were representative for the two volatile terpene classes. Photosynthetic characteristics partly recovered under moderate heat stress, and very limited recovery was observed under severe stress. All stress treatments resulted in elicitation of LOX emissions that declined during recovery. Enhanced mono- and sesquiterpene emissions were observed immediately after the heat treatment, but the emissions decreased even to below the control treatment during recovery between 2 and 10h, and raised again by 24h. The expression of β-phellandrene and (E)-β-caryophyllene synthase genes decreased between 2 and 10h after heat stress, and recovered to pre-stress level in mild heat stress treatment by 24h. Overall, this study demonstrates a highly sensitive heat response of terpenoid synthesis that is mainly controlled by gene level responses under mild stress, while severe stress leads to non-recoverable declines in foliage physiological and gene expression activities.

Role of isopentenyl-diphosphate isomerase in heterologous cyanobacterial (Synechocystis) isoprene production.

Heterologous production of isoprene (C5H8) hydrocarbons in cyanobacteria, emanating from sunlight, CO2, and water, is now attracting increasing attention. The concept entails application of an isoprene synthase transgene from terrestrial plants, heterologously expressed in cyanobacteria, aiming to reprogramcarbon flux in the terpenoid biosynthetic pathway toward formation and spontaneous release of this volatile chemical from the cell and liquid culture. However, flux manipulations and carbon-partitioning reactions between isoprene (the product) and native terpenoid biosynthesis for cellular needs are not yet optimized for isoprene yield. The primary reactant for isoprene biosynthesis is dimethylallyl diphosphate (DMAPP), whereas both DMAPP and its isopentenyl diphosphate (IPP) isomer are needed for cellular terpenoid biosynthesis. The present work addressed the function of an isopentenyl diphosphate (IPP) isomerase in cyanobacteria and its role in carbon partitioning between IPP and DMAPP, both of which serve, in variable ratios, as reactants for the synthesis of different cellular terpenoids. The work was approached upon the heterologous expression in Synechocystis of the "isopentenyl diphosphate isomerase" gene (FNI) from Streptococcus pneumoniae, using isoprene production as a "reporter process" for substrate partitioning between DMAPP and IPP. It is shown that transgenic expression of the FNI gene in Synechocystis resulted in a 250% increase in the "reporter isoprene" rate and yield, suggesting that the FNI isomerase shifted the endogenous DMAPP-IPP steady-state pool size toward DMAPP, thereby enhancing rates and yield of isoprene production. The work provides insight into the significance and functional role of the IPP isomerase in these photosynthetic microorganisms.

Temporal and spatial expression of terpene synthase genes associated with agarwood formation in Aquilaria malaccensis Lam

The diseased wood, agarwood, from the tropical tree taxa Aquilaria, is famed for its unique fragrance and medicinal values, mainly due to its richness in secondary metabolites such as the sesquiterpenes. The presence of sesquiterpenes in high numbers and amounts correlates with agarwood of high quality. To understand the synthesis of this important compound, we cloned two candidate genes in the terpenoid synthesis pathway from Aquilaria malaccensis Lam., a major agarwood tree species. The genes encoding sesquiterpene synthase (AmSesTPS1) and δ-guaiene synthase (AmGuaiS1) were successfully cloned from callus RNA, using specific primers derived from transcriptomic data, in a reverse transcription PCR reaction. The full-length complementary DNA (cDNA) sequence of AmSesTPS1 was 1632 bp encoding for 544 amino acids, and AmGuaiS1 was 1644 bp encoding for 547 amino acids. Sequence alignment analysis showed that AmSesTPS1 shared between 99 to 100 % identity with sesquiterpene synthase from Aquilaria sinensis (Lour.) Spreng. while AmGuaiS1 shared between 95 to 99 % identity with δ-guaiene synthases from Aquilaria crassna Pierre ex Lecomte and A. sinensis. The genes were functionally characterised in a time course wounding experiment using 3-year-old living trees. Two types of wood samples were collected: (1) from wounded area (S1) and (2) from 5 cm below the wounded area (S2). AmSesTPS1 was highly expressed after 6 h post wounding for both S1 and S2, at a level three- to six-fold higher than that of the unwounded control (0 h), while AmGuaiS1 was induced after just 2 h of wounding (18- and 5.5-fold at S1 and S2, respectively), after which the expression of both genes was lowered. The average normalised expression of both genes at S1 and S2 indicates the genes were also upregulated in the distal area from the wounding site. It can be deduced that wounding triggers these two genes in the sesquiterpene synthesis pathway, which ultimately leads to agarwood formation.

Rain-Shelter Cultivation Modifies Carbon Allocation in the Polyphenolic and Volatile Metabolism of Vitis vinifera L. Chardonnay Grapes.

This study investigated the effect of rain-shelter cultivation on the biosynthesis of flavonoids and volatiles in grapes, with an aim of determining whether rain-shelter application could help to improve the sensory attributes and quality of grapes. Vitis vinifera L. Chardonnay grapes, grown in the Huaizhuo basin region of northern China, were selected within two consecutive years. A rain-shelter roof was constructed using a colorless polyethylene (PE) film with a light transmittance of 80%. Results showed that rain-shelter treatment did not affect the accumulation of soluble solids during grape maturation. However, the allocation of assimilated carbon in phenolic and volatile biosynthetic pathways varied significantly, leading to alterations in polyphenolic and volatile profiles. The rain-shelter cultivation enhanced the concentration of flavan-3-ols via the flavonoid-3’5’-hydroxylase (F3’5’H) pathway, but reduced the level of flavonols and flavan-3-ols via the flavonoid-3’-hydroxylase (F3’H) pathway. In addition, the rain-shelter cultivation significantly enhanced the synthesis of fatty acid-derived volatiles, isoprene-derived terpenoids and amino acid-derived branched-chain aliphatics, but led to a decrease in the accumulation of isoprene-derived norisoprenoids and amino acid-derived benzenoids. Principal component analysis revealed some key compounds that differentiated the grapes cultivated under open-field and rain-shelter conditions. Moreover, the effect of the rain-shelter application on the accumulation of these compounds appeared to be vintage dependent. The alteration of their profiles caused by the rain-shelter treatment was significant in the vintage that received higher rainfall, which usually took place in the first rapid growth and veraison phases.

Reactive oxygen species and hormone signaling cascades in endophytic bacterium induced essential oil accumulation in Atractylodes lancea.

Pseudomonas fluorescens induces gibberellin and ethylene signaling via hydrogen peroxide in planta . Ethylene activates abscisic acid signaling. Hormones increase sesquiterpenoid biosynthesis gene expression and enzyme activity, inducing essential oil accumulation. Atractylodes lancea is a famous Chinese medicinal plant, whose main active components are essential oils. Wild A. lancea has become endangered due to habitat destruction and over-exploitation. Although cultivation can ensure production of the medicinal material, the essential oil content in cultivated A. lancea is significantly lower than that in the wild herb. The application of microbes as elicitors has become an effective strategy to increase essential oil accumulation in cultivated A. lancea. Our previous study identified an endophytic bacterium, Pseudomonas fluorescens ALEB7B, which can increase essential oil accumulation in A. lancea more efficiently than other endophytes; however, the underlying mechanisms remain unknown (Physiol Plantarum 153:30-42, 2015; Appl Environ Microb 82:1577-1585, 2016). This study demonstrates that P. fluorescens ALEB7B firstly induces hydrogen peroxide (H2O2) signaling in A. lancea, which then simultaneously activates gibberellin (GA) and ethylene (ET) signaling. Subsequently, ET activates abscisic acid (ABA) signaling. GA and ABA signaling increase expression of HMGR and DXR, which encode key enzymes involved in sesquiterpenoid biosynthesis, leading to increased levels of the corresponding enzymes and then an accumulation of essential oils. Specific reactive oxygen species and hormone signaling cascades induced by P. fluorescens ALEB7B may contribute to high-efficiency essential oil accumulation in A. lancea. Illustrating the regulation mechanisms underlying P. fluorescens ALEB7B-induced essential oil accumulation not only provides the theoretical basis for the inducible synthesis of terpenoids in many medicinal plants, but also further reveals the complex and diverse interactions among different plants and their endophytes.

Linking the response of endocrine regulated genes to adverse effects on sex differentiation improves comprehension of aromatase inhibition in a Fish Sexual Development Test

The Fish Sexual Development Test (FSDT) is a non-reproductive test to assess adverse effects of endocrine disrupting chemicals. With the present study it was intended to evaluate whether gene expression endpoints would serve as predictive markers of endocrine disruption in a FSDT. For proof-of-concept, a FSDT according to the OECD TG 234 was conducted with the non-steroidal aromatase inhibitor fadrozole (test concentrations: 10μg/L, 32μg/L, 100μg/L) using zebrafish (Danio rerio). Gene expression analyses using quantitative RT-PCR were included at 48h, 96h, 28days and 63days post fertilization (hpf, dpf). The selection of genes aimed at finding molecular endpoints which could be directly linked to the adverse apical effects of aromatase inhibition. The most prominent effects of fadrozole exposure on the sexual development of zebrafish were a complete sex ratio shift towards males and an acceleration of gonad maturation already at low fadrozole concentrations (10μg/L). Due to the specific inhibition of the aromatase enzyme (Cyp19) by fadrozole and thus, the conversion of C19-androgens to C18-estrogens, the steroid hormone balance controlling the sex ratio of zebrafish was altered. The resulting key event is the regulation of directly estrogen-responsive genes. Subsequently, gene expression of vitellogenin 1 (vtg1) and of the aromatase cyp19a1b isoform (cyp19a1b), were down-regulated upon fadrozole treatment compared to controls. For example, mRNA levels of vtg1 were down-regulated compared to the controls as early as 48 hpf and 96 hpf. Further regulated genes cumulated in pathways suggested to be controlled by endocrine mechanisms, like the steroid and terpenoid synthesis pathway (e.g. mevalonate (diphospho) decarboxylase (mvd), lanosterol synthase (2,3-oxidosqualene-lanosterol cyclase; lss), methylsterol monooxygenase 1 (sc4mol)) and in lipid transport/metabolic processes (steroidogenic acute regulatory protein (star), apolipoprotein Eb (apoEb)). Taken together, this study demonstrated that the existing Adverse Outcome Pathway (AOP) for aromatase inhibition in fish can be translated to the life-stage of sexual differentiation. We were further able to identify MoA-specific marker gene expression which can be instrumental in defining new measurable key events (KE) of existing or new AOPs related to endocrine disruption.

Laminarin modulates the chloroplast antioxidant system to enhance abiotic stress tolerance partially through the regulation of the defensin-like gene expression

Algae wall polysaccharide, laminarin (Lam), has an established role on induction of plant disease resistance. In this study, application of Lam increased Arabidopsis fresh weight and enhanced tolerance to salt and heat stress by stabilizing chloroplast under adverse environment. Transcriptome analysis indicated that, in addition to induced a large number of genes associated with the host defense, genes involved in the regulation of abiotic stress tolerance mostly the heat stress response constituted the largest group of the up-regulated genes. Lam induced expression of IRT1, ZIP8, and copper transporters involved in transport of Fe, Zn, Cu ions associated with the activity of chloroplast antioxidant system. Lam also up-regulated genes involved in the synthesis of terpenoid, a plastidial-derived secondary metabolite with antioxidant activity. Overexpression of a Lam-induced defensin like 202 (DEFL202) resulted in increased chloroplast stability under salt stress and increased plant growth activity after heat stress. Expression of antioxidant enzymes including SOD and ascorbate peroxidase (APX), photosystem PsbA-D1 and ABA-dependent responsive to desiccation 22 (RD22) was induced to higher levels in the transgenic seedlings. In sum, our results suggest that Lam is an potent inducer for induction of chloroplastic antioxidant activity. Lam affect plant abiotic stress tolerance partially through regulation of the DEFL-mediated pathway.

Trappc11 is required for protein glycosylation in zebrafish and humans.

Activation of the unfolded protein response (UPR) can be either adaptive or pathological. We term the pathological UPR that causes fatty liver disease a "stressed UPR." Here we investigate the mechanism of stressed UPR activation in zebrafish bearing a mutation in thetrappc11gene, which encodes a component of the transport protein particle (TRAPP) complex.trappc11mutants are characterized by secretory pathway defects, reflecting disruption of the TRAPP complex. In addition, we uncover a defect in protein glycosylation intrappc11mutants that is associated with reduced levels of lipid-linked oligosaccharides (LLOs) and compensatory up-regulation of genes in the terpenoid biosynthetic pathway that produces the LLO anchor dolichol. Treating wild-type larvae with terpenoid or LLO synthesis inhibitors phenocopies the stressed UPR seen intrappc11mutants and is synthetically lethal withtrappc11mutation. We propose that reduced LLO level causing hypoglycosylation is a mechanism of stressed UPR induction intrappc11mutants. Of importance, in human cells, depletion of TRAPPC11, but not other TRAPP components, causes protein hypoglycosylation, and lipid droplets accumulate in fibroblasts from patients with theTRAPPC11mutation. These data point to a previously unanticipated and conserved role for TRAPPC11 in LLO biosynthesis and protein glycosylation in addition to its established function in vesicle trafficking.

Proteomics of terpenoid biosynthesis and secretion in trichomes of higher plant species

Among the specialized (secondary) plant metabolites, terpenoids represent the most diverse family and are often involved in the defense against pathogens and herbivores. Terpenoids can be produced both constitutively and in response to the environment. At the front line of this defense strategy are the glandular trichomes, which are organs dedicated primarily to the production of specialized metabolites. Mass spectrometry-based proteomics is a powerful tool, which is very useful to investigate enzymes involved in metabolic pathways, such as the synthesis and secretion of terpenoids in glandular trichomes. Here we review the strategies used to investigate the specific roles of these particular organs from non-model plant species, mainly belonging to the Lamiaceae, Solanaceae, and Cannabaceae families. We discuss how proteomics helps to accurately pinpoint candidate proteins to be functionally characterized, and how technological progresses create opportunities for studying low-abundance proteins, such as the ones related to the synthesis and transport of specialized metabolites. This article is part of a Special Issue entitled: Plant Proteomics — a bridge between fundamental processes and crop production, edited by Dr. Hans-Peter Mock.

The genome of the seagrass Zostera marina reveals angiosperm adaptation to the sea.

Seagrasses colonized the sea on at least three independent occasions to form the basis of one of the most productive and widespread coastal ecosystems on the planet. Here we report the genome of Zostera marina (L.), the first, to our knowledge, marine angiosperm to be fully sequenced. This reveals unique insights into the genomic losses and gains involved in achieving the structural and physiological adaptations required for its marine lifestyle, arguably the most severe habitat shift ever accomplished by flowering plants. Key angiosperm innovations that were lost include the entire repertoire of stomatal genes, genes involved in the synthesis of terpenoids and ethylene signalling, and genes for ultraviolet protection and phytochromes for far-red sensing. Seagrasses have also regained functions enabling them to adjust to full salinity. Their cell walls contain all of the polysaccharides typical of land plants, but also contain polyanionic, low-methylated pectins and sulfated galactans, a feature shared with the cell walls of all macroalgae and that is important for ion homoeostasis, nutrient uptake and O2/CO2 exchange through leaf epidermal cells. The Z. marina genome resource will markedly advance a wide range of functional ecological studies from adaptation of marine ecosystems under climate warming, to unravelling the mechanisms of osmoregulation under high salinities that may further inform our understanding of the evolution of salt tolerance in crop plants.

Endophytic Bacterium-Triggered Reactive Oxygen Species Directly Increase Oxygenous Sesquiterpenoid Content and Diversity in Atractylodes lancea.

Oxygenous terpenoids are active components of many medicinal plants. However, current studies that have focused on enzymatic oxidation reactions cannot comprehensively clarify the mechanisms of oxygenous terpenoid synthesis and diversity. This study shows that an endophytic bacterium can trigger the generation of reactive oxygen species (ROS) that directly increase oxygenous sesquiterpenoid content and diversity in Atractylodes lancea. A. lancea is a famous but endangered Chinese medicinal plant that contains abundant oxygenous sesquiterpenoids. Geo-authentic A. lancea produces a wider range and a greater abundance of oxygenous sesquiterpenoids than the cultivated herb. Our previous studies have shown the mechanisms behind endophytic promotion of the production of sesquiterpenoid hydrocarbon scaffolds; however, how endophytes promote the formation of oxygenous sesquiterpenoids and their diversity is unclear. After colonization by Pseudomonas fluorescens ALEB7B, oxidative burst and oxygenous sesquiterpenoid accumulation in A. lancea occur synchronously. Treatment with exogenous hydrogen peroxide (H2O2) or singlet oxygen induces oxidative burst and promotes oxygenous sesquiterpenoid accumulation in planta. Conversely, pretreatment of plantlets with the ROS scavenger ascorbic acid significantly inhibits the oxidative burst and oxygenous sesquiterpenoid accumulation induced by P. fluorescens ALEB7B. Further in vitro oxidation experiments show that several oxygenous sesquiterpenoids can be obtained from direct oxidation caused by H2O2 or singlet oxygen. In summary, this study demonstrates that endophytic bacterium-triggered ROS can directly oxidize oxygen-free sesquiterpenoids and increase the oxygenous sesquiterpenoid content and diversity in A. lancea, providing a novel explanation of the mechanisms of oxygenous terpenoid synthesis in planta and an essential complementarity to enzymatic oxidation reactions.

Cloning and expression analysis of 4-(cytidine-5-diphospho)-2-C-methyl-D-erythritol kinase gene in Tripterygium wilfordii

4-(Cytidine-5-diphospho) -2-C-methyl-D-erythritol kinase is a key enzyme in the biosynthesis pathway of terpenoids. According to the transcriptome database, the specific primers were designed and used in PCR. The bioinformatic analysis of the sequenced TwCMK gene was performed in several bioinformatics software. The Real-time fluorescence quantification polymerase chain reaction (RT-qPCR) were used to detect the expression levels of TwCMK from T. wilfordii after elicitor MeJA supplied. The results showed that the full length of TwCMK cDNA was 1 732 bp encoding 387 amino acids. The theoretical isoelectric point of the putative TwCMK protein was 5.79 and the molecular weight was about 42.85 kDa. MeJA stimulated the rising of TwCMK expression in suspension cell and signally impacted at 24 h. The research provides a basis for further study on the regulation of terpenoid secondary metabolism and biological synthesis.

Bioconversion of methanol to value-added mevalonate by engineered Methylobacterium extorquens AM1 containing an optimized mevalonate pathway.

Methylotrophic biosynthesis using methanol as a feedstock is a promising and attractive method to solve the over-dependence of the bioindustry on sugar feedstocks derived from grains that are used for food. In this study, we introduced and engineered the mevalonate pathway into Methylobacterium extorquens AM1 to achieve high mevalonate production from methanol, which could be a platform for terpenoid synthesis. We first constructed a natural operon (MVE) harboring the mvaS and mvaE genes from Enterococcus faecalis as well as an artificial operon (MVH) harboring the hmgcs1 gene from Blattella germanica and the tchmgr gene from Trypanosoma cruzi that encoded enzymes with the highest reported activities. We achieved mevalonate titers of 56 and 66 mg/L, respectively, in flask cultivation. Introduction of the phaA gene from Ralstonia eutropha into the operon MVH increased the mevalonate titer to 180 mg/L, 3.2-fold higher than that of the natural operon MVE. Further modification of the expression level of the phaA gene by regulating the strength of the ribosomal binding site resulted in an additional 20 % increase in mevalonate production to 215 mg/L. A fed-batch fermentation of the best-engineered strain yielded a mevalonate titer of 2.22 g/L, which was equivalent to an overall yield and productivity of 28.4 mg mevalonate/g methanol and 7.16 mg/L/h, respectively. The production of mevalonate from methanol, which is the initial, but critical step linking methanol with valuable terpenoids via methylotrophic biosynthesis, represents a proof of concept for pathway engineering in M. extorquens AM1.

Engineering Methylobacterium extorquens for de novo synthesis of the sesquiterpenoid α-humulene from methanol

Over the last 10 to 15 years, metabolic engineering of microbes has become a versatile tool for high-level de novo synthesis of terpenoids, with the sesquiterpenoids armopha-1,4-diene, farnesene and artemisinic acid as prime examples. However, almost all cell factory approaches towards terpenoids to date have been based on sugar as the raw material, which is mainly used as a food resource and subject to high price volatilities. In this study we present de novo synthesis of the sesquiterpenoid α-humulene from the abundantly available non-food carbon source methanol by metabolically engineered Methylobacterium extorquens AM1. Expression of α-humulene synthase from Zingiber zerumbet in combination with farnesyl pyrophosphate (FPP) synthase from Saccharomyces cerevisiae led to concentrations of up to 18mg/L α-humulene. Introduction of a prokaryotic mevalonate pathway from Myxococcus xanthus in combination with ribosome binding site optimization of α-humulene and FPP synthases increased product concentration 3-fold. This value was additionally raised by 30% using a carotenoid synthesis deficient mutant strain. Final product concentrations of up to 1.65g/L were obtained in methanol limited fed-batch cultivations, which is the highest titer of de novo synthesized α-humulene reported to date. This study demonstrates the potential of M. extorquens as a future platform strain for the production of high-value terpenoids from the alternative carbon source methanol.