Abstract
Isoprene, a Natural Volatile Organic Compound (NVOC) is one of the chief by-products of plant metabolism with important applications in the synthesis of rubber and pharmaceuticals as a platform molecule. Isoprene was obtained earlier from petroleum sources; however, to synthesise it new fermentation-based strategies are being adopted. Bioinformatics tools were utilised to isolate the Isoprene Synthase (IspS) gene which converts the precursors Isopentenyl Diphosphate (IPP) and Dimethylallyl Diphosphate (DMAPP) into isoprene. Metabolic engineering strategies were to synthesise an isoprene-producing recombinant clone derived from Artocarpus heterophyllus (jackfruit). The functional characterization was done using the overexpression of the isoprene synthase gene in an Escherichia coli BL21 host. The recombinant clone, ISPS_GBL_001 (submitted to GenBank, National Centre for Biotechnology Information or NCBI) was used for fermentation in the batch and fed-batch mode to produce isoprene. Isoprene productivity of 0.08 g/g dextrose was obtained via the fed-batch mode maintaining the process parameters at optimum. The quantification and confirmation of isoprene was done using gas chromatography (GC) and GC-mass spectrometry (GC–MS) of the extracted sample, respectively. This study makes significant contribution to the ongoing research on bio-isoprene synthesis by highlighting a novel plant source of the IspS gene followed by, its successful expression in a recombinant host, validated by fermentation.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
The data that supports the findings of this study are available within the article and its supplementary material.
References
Abdul Rahman SN, Bakar MFA, Singham GV, Othman AS (2019) Single-nucleotide polymorphism markers within MVA and MEP pathways among Hevea brasiliensis clones through transcriptomic analysis. 3 Biotech 9:1–10. https://doi.org/10.1007/s13205-019-1921-3
Abraham OSJ, Miguel TS, Inocencio HC, Blondy CC (2017) A quick and effective in-house method of DNA purification from agarose gel, suitable for sequencing. 3 Biotech. https://doi.org/10.1007/s13205-017-0851-1
Alam MI, Gupta S, Ahmad E, Haider MA (2015) Integrated bio- and chemocatalytic processing for biorenewable chemicals and fuels. Sustainable catalytic processes. Elsevier
Batten R, Karanjikar M, Spatari S (2021) Bio-based polyisoprene can mitigate climate change and deforestation in expanding rubber production. Fermentation 7:1–14. https://doi.org/10.3390/fermentation7040204
Chaliyakunnel S, Millet DB, Chen X (2019) Constraining emissions of volatile organic compounds over the indian subcontinent using space-based formaldehyde measurements. J Geophys Res Atmos 124:10525–10545. https://doi.org/10.1029/2019JD031262
Chang E, Zhao Y, Wei Q et al (2016) Isolation of high-quality RNA from Platycladus orientalis and other Cupressaceae plants. Electron J Biotechnol 23:21–27. https://doi.org/10.1016/j.ejbt.2016.08.003
Fels U, Gevaert K, Van Damme P (2020) Bacterial genetic engineering by means of recombineering for reverse genetics. Front Microbiol 11:548410
Freeman LA (2013) Cloning full-length transcripts and transcript variants using 5′ and 3′ RACE. Methods Mol Biol 1027:3–17. https://doi.org/10.1007/978-1-60327-369-5_1
Gomaa L, Loscar ME, Zein HS et al (2017a) Boosting isoprene production via heterologous expression of the Kudzu isoprene synthase gene (kIspS) into Bacillus spp. cell factory. AMB Express. https://doi.org/10.1186/s13568-017-0461-7
Gomaa L, Zein H, Abdel Ghaffar N, Abdallah N (2017b) Molecular studies on isoprene production from natural and recombinant bacteria. J Agric Chem Biotechnol 8:65–69. https://doi.org/10.21608/jacb.2017.38435
Guo J, Huang Z, Sun J et al (2021) Research progress and future development trends in medicinal plant transcriptomics. Front Plant Sci 12:1–10. https://doi.org/10.3389/fpls.2021.691838
Harvey CM, Sharkey TD (2016) Exogenous isoprene modulates gene expression in unstressed Arabidopsis thaliana plants. Plant Cell Environ 39:1251–1263. https://doi.org/10.1111/pce.12660
Himmelberger JA, Cole KE, Dowling DP (2018) Biocatalysis: nature’s chemical toolbox. Green Chem Incl Approach. https://doi.org/10.1016/B978-0-12-809270-5.00019-4
Jing X, Lun X, Fan C, Ma W (2020) Emission patterns of biogenic volatile organic compounds from dominant forest species in Beijing, China. J Environ Sci (china) 95:73–81. https://doi.org/10.1016/j.jes.2020.03.049
Kim JH, Wang C, Jang HJ et al (2016) Isoprene production by Escherichia coli through the exogenous mevalonate pathway with reduced formation of fermentation byproducts. Microb Cell Fact. https://doi.org/10.1186/s12934-016-0612-6
Koppolu V, Vasigala VK (2016) Role of Escherichia coli in biofuel production. Microbiol Insights. https://doi.org/10.4137/mbi.s10878
Lantz AT, Allman J, Weraduwage SM, Sharkey TD (2019) Control of rate and physiological role of isoprene emission from plants. Plant Cell Environ 42:13629
Lee HW, Park JH, Lee HS et al (2019) Production of bio-based isoprene by the mevalonate pathway cassette in Ralstonia eutropha. J Microbiol Biotechnol 29:1656–1664. https://doi.org/10.4014/jmb.1909.09002
Lee HW, Park JH, Kim WK et al (2020) Engineered Escherichia coli strains as platforms for biological production of isoprene. FEBS Open Biol 10:780–788. https://doi.org/10.1002/2211-5463.12829
Li M, Nian R, Xian M, Zhang H (2018) Metabolic engineering for the production of isoprene and isopentenol by Escherichia coli. Appl Microbiol Biotechnol 102:7725–7738. https://doi.org/10.1007/s00253-018-9200-5
Liu L, Han R, Yu N et al (2018) A method for extracting high-quality total rna from plant rich in polysaccharides and polyphenols using dendrobium huoshanense in researching the key technologies concerning the integration of processing Chinese medicine decoction pieces in producing area. PLoS ONE. https://doi.org/10.1371/journal.pone.0196592
Makam S, Srirama K, Dirisala VR, Reddy PN (2018) An efficient method for integration of PCR fragments into adjacent or overlapping restriction sites during gene cloning. 3 Biotech. https://doi.org/10.1007/s13205-018-1214-2
Martin VJJ, Piteral DJ, Withers ST et al (2003) Engineering a mevalonate pathway in Escherichia coli for production of terpenoids. Nat Biotechnol 21:796–802. https://doi.org/10.1038/nbt833
Mcgenity TJ, Crombie AT, Murrell CJ (2018) Microbial cycling of isoprene, the most abundantly produced biological volatile organic compound on earth background and isoprene production. ISME J 12:931–941. https://doi.org/10.1038/s41396-018-0072-6
Moe KT, Chung JW, Il CY et al (2011) Sequence information on simple sequence repeats and single nucleotide polymorphisms through transcriptome analysis of mungbean. J Integr Plant Biol 53:63–73. https://doi.org/10.1111/j.1744-7909.2010.01012.x
Morais ARC, Dworakowska S, Reis A et al (2015) Chemical and biological-based isoprene production: green metrics. Catal Today 239:38–43. https://doi.org/10.1016/j.cattod.2014.05.033
Nitta N, Tajima Y, Katashkina JI et al (2020) Application of inorganic phosphate limitation to efficient isoprene production in Pantoea ananatis. J Appl Microbiol 128:763–774. https://doi.org/10.1111/jam.14521
Pade N, Erdmann S, Enke H et al (2016) Insights into isoprene production using the cyanobacterium Synechocystis sp. PCC 6803. Biotechnol Biofuels 9:1–16. https://doi.org/10.1186/s13068-016-0503-4
Padmanabhan P, Srikakulam N, Velayudha Vimala Kumar K et al (2018) Comprehensive leaf transcriptome of a non-model plant, Abelmoschus esculentus for the functional genomics studies. J Genet Genome Res 5:1–14. https://doi.org/10.23937/2378-3648/1410036
Patel AA, Shukla YM, Kumar S et al (2020) Transcriptome analysis for molecular landscaping of genes controlling diterpene andrographolide biosynthesis in Andrographis paniculata (Burm. f) Nees. 3. Biotech 10:1–14. https://doi.org/10.1007/s13205-020-02511-y
Sanadze GA (2004) Biogenic isoprene (a review). Russ J Plant Physiol 51:729–741. https://doi.org/10.1023/B:RUPP.0000047821.63354.a4
Sharkey TD, Monson RK (2017) Isoprene research: 60 years later, the biology is still enigmatic. Plant Cell Environ 40:1671–1678. https://doi.org/10.1111/pce.12930
Silver GM, Fall R (1995) Characterization of aspen isoprene synthase, an enzyme responsible for leaf isoprene emission to the atmosphere. J Biol Chem 270:13010–13016. https://doi.org/10.1074/jbc.270.22.13010
Singh B, Sharma RA (2015) Plant terpenes: defense responses, phylogenetic analysis, regulation and clinical applications. 3 Biotech 5:129–151. https://doi.org/10.1007/s13205-014-0220-2
Singh R, Singh AP, Singh MP et al (2008) Emission of isoprene from common Indian plant species and its implications for regional air quality. Environ Monit Assess 144:43–51. https://doi.org/10.1007/s10661-007-9940-7
Stadler BM, Wulf C, Werner T et al (2019) Catalytic approaches to monomers for polymers based on renewables. ACS Catal 9:8012–8067. https://doi.org/10.1021/acscatal.9b01665
Varshney CK, Singh AP (2003) Isoprene emission from Indian trees. J Geophys Res Atmos. https://doi.org/10.1029/2003jd003866
Volke DC, Rohwer J, Fischer R, Jennewein S (2019) Investigation of the methylerythritol 4-phosphate pathway for microbial terpenoid production through metabolic control analysis. Microb Cell Fact 18:1–15. https://doi.org/10.1186/s12934-019-1235-5
Wittmann C, Liao JC (2017) Industrial biotechnology: products and processes. In: Wittmann. C, Liao JC (eds) Part VI top-value platform chemicals. Wiley-VCH Verlag GmbH \& Co. KGaA, Germany, pp. 475–504. https://www.wiley.com/en-us/Industrial+Biotechnology%3A+Products+and+Processes-p-9783527341818
Xue J, Ahring BK (2011) Enhancing isoprene production by genetic modification of the 1-deoxy-D-Xylulose-5-phosphate pathway in Bacillus subtilis. Appl Environ Microbiol 77:2399–2405. https://doi.org/10.1128/AEM.02341-10
Yang J, Nie Q, Liu H et al (2016) A novel MVA-mediated pathway for isoprene production in engineered E. coli. BMC Biotechnol 16:1–9. https://doi.org/10.1186/s12896-016-0236-2
Ye L, Lv X, Yu H (2016) Engineering microbes for isoprene production. Metab Eng 38:125–138. https://doi.org/10.1016/j.ymben.2016.07.005
Yeom SJ, Kim M, Kim SK et al (2018) Molecular and biochemical characterization of a novel isoprene synthase from Metrosideros polymorpha. BMC Plant Biol 18:1–10. https://doi.org/10.1186/s12870-018-1315-4
Zhao C, Zhao XY, Liu JJ et al (2018) Enhanced D-ribose production by genetic modification and medium optimization in Bacillus subtilis 168. Korean J Chem Eng 35:1137–1143. https://doi.org/10.1007/s11814-017-0356-y
Zhou J, Lu X, Tian B et al (2020) A gas chromatography-flame ionization detection method for direct and rapid determination of small molecule volatile organic compounds in aqueous phase. 3 Biotech 10:1–9. https://doi.org/10.1007/s13205-020-02523-8
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors declare there were no conflicts of interest.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
About this article
Cite this article
Dive, A., Singhal, R., Srivastava, S. et al. Isolation and functional characterization of novel isoprene synthase from Artocarpus heterophyllus (jackfruit). 3 Biotech 13, 24 (2023). https://doi.org/10.1007/s13205-022-03441-7
Received:
Accepted:
Published:
DOI: https://doi.org/10.1007/s13205-022-03441-7