Production of 24- Methylenecholesterol from Royal Jelly by <i>Nannocloropsis oceanica</i> IMET1

doi:10.13523/j.cb.2204070

China Biotechnology

2022, Vol. 42

Issue (8): 30-39 DOI: 10.13523/j.cb.2204070

Production of 24- Methylenecholesterol from Royal Jelly by Nannocloropsis oceanica IMET1

DENG Xiang-zi,ZHOU Wen-xu,LU Yan-du^**(

)

State Key Laboratory of Marine Resource Utilization in South China Sea, College of Oceanology, Hainan University, Haikou 570228, China

Download:

HTML

PDF(3527KB) HTML
Export: BibTeX | EndNote (RIS)

Abstract

Microalgal biotechnology for biofuels is at a crossroads and its development is still in flux. The development of microalga-derived multi-product technology will greatly improve the economic viability, particularly with a combination of the production of high value-added compounds. The results showed that the mutants with DWARF1 (DWF1) gene knocked out had higher pigment content and photosynthetic efficiency than the wild type, and could also significantly reduce the accumulation of cholesterol (which might act as a leading risk factor for human cardiovascular disease) from over 70% of total sterols (TSs) to null. In contrast, the production of its precursor 24-methylenecholesterol (a critical micronutrient of royal jelly that is beneficial to human health) was increased from null to more than 60% of TSs. Combined with the high content of omega-3 fatty acids of N. oceanica, we anticipate an appreciable profit by exploiting this strain on an industrial scale.

Key words： Nannocloropsis oceanica DWARF1 gene Cholesterol 24-Methylenecholesterol Phytosterols Honeybee artificial food Royal jelly

Received: 27 April 2022 Published: 07 September 2022

ZTFLH:

Q819

Corresponding Authors: Yan-du LU E-mail: ydlu@hainanu.edu.cn

	Service
	E-mail this article
	Add to my bookshelf
	Add to citation manager
	E-mail Alert
	RSS
	Articles by authors
	DENG Xiang-zi
	ZHOU Wen-xu
	LU Yan-du

Cite this article:

DENG Xiang-zi, ZHOU Wen-xu, LU Yan-du. Production of 24- Methylenecholesterol from Royal Jelly by Nannocloropsis oceanica IMET1. China Biotechnology, 2022, 42(8): 30-39.

URL:

https://manu60.magtech.com.cn/biotech/10.13523/j.cb.2204070 OR https://manu60.magtech.com.cn/biotech/Y2022/V42/I8/30

Fig.1 PCR amplification of the genomic DNA of the dwf1 mutants M:DL2000;Lanes1-9:The samples of the dwf1 mutants;NC:Negative control

Fig.2 Growth performance of the dwf1 mutants and wild type N. oceanic (WT) under the normal condition (a) Optical density at 750 nm of the dwf1 mutants and WT (b) Growth curve of the dwf1 mutants and WT (c) Pigment content in per unit volume of the dwf1 mutants and WT (d) Pigment content in per cell of the dwf1 mutants and the WT (e) Dry weight in the dwf1 mutants and the WT (f) Maximum quantum yield of photosystem II measurements of the dwf1 mutants and the WT Cultured under 25℃, 50 μmol photons/(m²·s) light intensity. Data are presentedas means ± SDs (n=3). Asterisks (*) indicate statistically significant differences between WT and the mutants in designated conditions (P< 0.05)

Table 1 GC-MS data for sterol profiling from WT and dwf1 mutants under the normal condition

Fig.S1 Genome sequence of the wild type N.oceanica

Fig.S2 Genome sequence of the dwf1-1 mutant

Fig.S3 Genome sequence of the dwf1-7 mutant

Fig.S4 The main sterol kinds of N. oceanica


[1]	Rodolfi L, Chini Zittelli G, Bassi N, et al. Microalgae for oil: strain selection, induction of lipid synthesis and outdoor mass cultivation in a low-cost photobioreactor. Biotechnology and Bioengineering, 2009, 102(1): 100-112. doi: 10.1002/bit.22033

[2]	Chen C Y, Chen Y C, Huang H C, et al. Engineering strategies for enhancing the production of eicosapentaenoic acid (EPA) from an isolated microalga Nannochloropsis oceanica CY2. Bioresource Technology, 2013, 147: 160-167. doi: 10.1016/j.biortech.2013.08.051

[3]	Llaverias G, Escolà-Gil J C, Lerma E, et al. Phytosterols inhibit the tumor growth and lipoprotein oxidizability induced by a high-fat diet in mice with inherited breast cancer. The Journal of Nutritional Biochemistry, 2013, 24(1): 39-48. doi: 10.1016/j.jnutbio.2012.01.007

[4]	Escurriol V, Cofán M, Moreno-Iribas C, et al. Phytosterol plasma concentrations and coronary heart disease in the prospective Spanish EPIC cohort. Journal of Lipid Research, 2010, 51(3): 618-624. doi: 10.1194/jlr.P000471 pmid: 19786566

[5]	Demonty I, Ras R T, van der Knaap H C M, et al. The effect of plant sterols on serum triglyceride concentrations is dependent on baseline concentrations: a pooled analysis of 12 randomised controlled trials. European Journal of Nutrition, 2013, 52(1): 153-160. doi: 10.1007/s00394-011-0297-x

[6]	Lye C T, Mukherjee S, Burns S F. Combining plant sterols with walking lowers postprandial triacylglycerol more than walking only in Chinese men with elevated body mass index. International Journal of Sport Nutrition and Exercise Metabolism, 2019, 29(6): 576-582. doi: 10.1123/ijsnem.2018-0398

[7]	Trautwein E A, Koppenol W P, de Jong A, et al. Plant sterols lower LDL-cholesterol and triglycerides in dyslipidemic individuals with or at risk of developing type 2 diabetes; a randomized, double-blind, placebo-controlled study. Nutrition & Diabetes, 2018, 8(1): 30.

[8]	郭盼盼, 韩婷. 植物甾醇与心血管疾病关系的研究进展. 中国食物与营养, 2021, 27(9): 5-8.

[8]	Guo P P, Han T. Research advancement on the relationship between phytosterol and cardiovascular disease. Food and Nutrition in China, 2021, 27(9): 5-8.

[9]	Moll P P, Sing C F, Weidman W H, et al. Total cholesterol and lipoproteins in school children: prediction of coronary heart disease in adult relatives. Circulation, 1983, 67(1): 127-134. pmid: 6847791

[10]	McGill H C Jr, McMahan C A, Gidding S S. Preventing heart disease in the 21st century: implications of the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study. Circulation, 2008, 117(9): 1216-1227. doi: 10.1161/CIRCULATIONAHA.107.717033 pmid: 18316498

[11]	Wu Q Q, Wang Q T, Fu J F, et al. Polysaccharides derived from natural sources regulate triglyceride and cholesterol metabolism: a review of the mechanisms. Food & Function, 2019, 10(5): 2330-2339.

[12]	le Goff M, le Ferrec E, Mayer C, et al. Microalgal carotenoids and phytosterols regulate biochemical mechanisms involved in human health and disease prevention. Biochimie, 2019, 167: 106-118. doi: 10.1016/j.biochi.2019.09.012

[13]	Lu Y D, Zhou W X, Wei L, et al. Regulation of the cholesterol biosynthetic pathway and its integration with fatty acid biosynthesis in the oleaginous microalga Nannochloropsis oceanica. Biotechnology for Biofuels, 2014, 7: 81. doi: 10.1186/1754-6834-7-81

[14]	Pain J, Barbier M, Bogdanovsky D, et al. Chemistry and biological activity of the secretions of queen and worker honeybees (Apis mellifica L.). Comparative Biochemistry and Physiology, 1962, 6(3): 233-241. doi: 10.1016/0010-406X(62)90081-6

[15]	Brown W H, Felauer E E, Freure R J. Some new components of royal jelly. Canadian Journal of Chemistry, 1961, 39(5): 1086-1089. doi: 10.1139/v61-134

[16]	Herbert E W Jr, Svoboda J A, Thompson M J, et al. Sterol utilization in honey bees fed a synthetic diet: effects on brood rearing. Journal of Insect Physiology, 1980, 26(5): 287-289. doi: 10.1016/0022-1910(80)90135-3

[17]	Chakrabarti P, Lucas H M, Sagili R R. Evaluating effects of a critical micronutrient (24-methylenecholesterol) on honey bee physiology. Annals of the Entomological Society of America, 2019, 113(3): 176-182. doi: 10.1093/aesa/saz067

[18]	Sugiyama T, Takahashi K, Tokoro S, et al. Inhibitory effect of 10-hydroxy-trans-2-decenoic acid on LPS-induced IL-6 production via reducing IκB-ζ expression. Innate Immunity, 2012, 18(3): 429-437. doi: 10.1177/1753425911416022

[19]	Park M J, Kim B Y, Park H G, et al. Major royal jelly protein 2 acts as an antimicrobial agent and antioxidant in royal jelly. Journal of Asia-Pacific Entomology, 2019, 22(3): 684-689. doi: 10.1016/j.aspen.2019.05.003

[20]	Jiang C M, Liu X, Li C X, et al. Anti-senescence effect and molecular mechanism of the major royal jelly proteins on human embryonic lung fibroblast (HFL-I) cell line. Journal of Zhejiang University Science B, 2018, 19(12): 960-972. doi: 10.1631/jzus.B1800257

[21]	Pan Y M, Rong Y L, You M M, et al. Royal jelly causes hypotension and vasodilation induced by increasing nitric oxide production. Food Science & Nutrition, 2019, 7(4): 1361-1370.

[22]	Guardia de Souza e Silva T, do Val de Paulo M E F, da Silva J R M, et al. Oral treatment with royal jelly improves memory and presents neuroprotective effects on icv-STZ rat model of sporadic Alzheimer’s disease. Heliyon, 2020, 6(2): e03281. doi: 10.1016/j.heliyon.2020.e03281

[23]	Bincoletto C, Eberlin S, Figueiredo C A V, et al. Effects produced by Royal Jelly on haematopoiesis: relation with host resistance against Ehrlich ascites tumour challenge. International Immunopharmacology, 2005, 5(4): 679-688. doi: 10.1016/j.intimp.2004.11.015 pmid: 15710337

[24]	Lu Y D, Tarkowská D, Turečková V, et al. Antagonistic roles of abscisic acid and cytokinin during response to nitrogen depletion in oleaginous microalga Nannochloropsis oceanica expand the evolutionary breadth of phytohormone function. The Plant Journal, 2014, 80(1): 52-68. doi: 10.1111/tpj.12615

[25]	Poliner E, Takeuchi T, Du Z Y, et al. Nontransgenic marker-free gene disruption by an episomal CRISPR system in the oleaginous microalga, Nannochloropsis oceanica CCMP1779. ACS Synthetic Biology, 2018, 7(4): 962-968. doi: 10.1021/acssynbio.7b00362 pmid: 29518315

[26]	Lu Y D, Gan Q H, Iwai M, et al. Role of an ancient light-harvesting protein of PSI in light absorption and photoprotection. Nature Communications, 2021, 12: 679. doi: 10.1038/s41467-021-20967-1

[27]	Wang Q T, Lu Y D, Xin Y, et al. Genome editing of model oleaginous microalgae Nannochloropsis spp. by CRISPR/Cas9. The Plant Journal, 2016, 88(6): 1071-1081. doi: 10.1111/tpj.13307

[28]	Zhou W X, Branch W D, Gilliam L, et al. Phytosterol composition of Arachis hypogaea seeds from different maturity classes. Molecules (Basel, Switzerland), 2018, 24(1): 106. doi: 10.3390/molecules24010106

[29]	Zhou W X, Cross G A M, Nes W D. Cholesterol import fails to prevent catalyst-based inhibition of ergosterol synthesis and cell proliferation of Trypanosoma brucei. Journal of Lipid Research, 2007, 48(3): 665-673. doi: 10.1194/jlr.M600404-JLR200

[30]	Takahashi T, Gasch A, Nishizawa N, et al. The DIMINUTO gene of Arabidopsis is involved in regulating cell elongation. Genes & Development, 1995, 9(1): 97-107. doi: 10.1101/gad.9.1.97

[31]	Klahre U, Noguchi T, Fujioka S, et al. The Arabidopsis DIMINUTO/DWARF1 gene encodes a protein involved in steroid synthesis. The Plant Cell, 1998, 10(10): 1677-1690. doi: 10.1105/tpc.10.10.1677

[32]	Choe S, Dilkes B P, Gregory B D, et al. The Arabidopsis dwarf 1 mutant is defective in the conversion of 24-methylenecholesterol to campesterol in brassinosteroid biosynthesis. Plant Physiology, 1999, 119(3): 897-908. pmid: 10069828

[33]	Tsukagoshi Y, Suzuki H, Seki H, et al. Ajuga Δ24-sterol reductase catalyzes the direct reductive conversion of 24-methylenecholesterol to campesterol. The Journal of Biological Chemistry, 2016, 291(15): 8189-8198. doi: 10.1074/jbc.M115.703470

[34]	Youn J H, Kim T W, Joo S H, et al. Function and molecular regulation of DWARF 1 as a C-24 reductase in brassinosteroid biosynthesis in Arabidopsis. Journal of Experimental Botany, 2018, 69(8): 1873-1886. doi: 10.1093/jxb/ery038

[35]	Du L Q, Poovaiah B W. Ca2+/calmodulin is critical for brassinosteroid biosynthesis and plant growth. Nature, 2005, 437(7059): 741-745. doi: 10.1038/nature03973

[36]	Ajjawi I, Verruto J, Aqui M, et al. Lipid production in Nannochloropsis gaditana is doubled by decreasing expression of a single transcriptional regulator. Nature Biotechnology, 2017, 35(7): 647-652. doi: 10.1038/nbt.3865 pmid: 28628130

[37]	Kang N K, Kim E K, Kim Y U, et al. Increased lipid production by heterologous expression of AtWRI 1 transcription factor in Nannochloropsis salina. Biotechnology for Biofuels, 2017, 10: 231. doi: 10.1186/s13068-017-0919-5 pmid: 29046718

[38]	Han X, Song X J, Li F L, et al. Improving lipid productivity by engineering a control-knob gene in the oleaginous microalga Nannochloropsis oceanica. Metabolic Engineering Communications, 2020, 11: e00142. doi: 10.1016/j.mec.2020.e00142

[39]	Yang F, Yuan W, Ma Y H, et al. Harnessing the lipogenic potential of Δ6-desaturase for simultaneous hyperaccumulation of lipids and polyunsaturated fatty acids in Nannochloropsis oceanica. Frontiers in Marine Science, 2019, 6: 682. doi: 10.3389/fmars.2019.00682

[1]	Qian-qian GUO,Deng-ke GAO,Xiao-tao CHENG,Fu-ping LU,Tanokura Masaru,Hui-min QIN. Heterologous Expression, Purification and Enzymatic Characterization of Cholesterol Oxidase PsCO₄[J]. China Biotechnology, 2018, 38(6): 34-42.

[2]	ZHANG Yu-fu, WANG Jian-wen, LI Song-tao, ZHU Zhang-liang, LU Fu-ping, MAO Shu-hong, QIN Hui-min. The Expression, Purification and Structural Analysis of Cholesterol Oxidase ChOG[J]. China Biotechnology, 2017, 37(6): 43-49.

[3]	ZHANG Jing, ZHANG Wen-qiang, QIN Hui-min, MAO Shu-hong, XUE Jia-lu, LU Fu-ping. Expression of Related Cholesterol Dehydrogenation Protein and Bioactivity Research[J]. China Biotechnology, 2017, 37(1): 21-26.

[4]	ZHANG Wen-qian, XIAO Wen-hai, ZHOU Xiao, WANG Ying. Effect of Post-squalene Genes on the Synthesis of 7-Dehydrocholesterol in the Artificial Saccharomyces cerevisiae[J]. China Biotechnology, 2016, 36(6): 39-50.

[5]	ZHANG Jing, ZHANG Yu fu, QIN Hui min, MAO Shu hong, LU Fu ping. *Construction of Artificial Arthrobacter simplex* for Oxidizing Cholesterol, and Optimization of Bioconversion Condition**[J]. China Biotechnology, 2016, 36(11): 70-75.

[6]	. Establish an ELISA Method for Screening Agonists of the Rattus Norvegicus LXRβ[J]. China Biotechnology, 2010, 30(07): 0-0.

Viewed

Full text

Abstract

Cited

Shared

Discussed