Abstract
Main conclusion
This study showed that a galactosyltransferase, AgUCGalT1, is involved in anthocyanin galactosylation in purple celery.
Celery is a well-known vegetable because of its rich nutrients, low calories, and medicinal values. Its petioles and leaf blades are the main organs acting as nutrient sources. UDP-galactose: cyanidin 3-O-galactosyltransferase can transfer the galactosyl moiety from UDP-galactose to the 3-O-position of cyanidin through glycosylation. This process enhances the stability and water solubility of anthocyanins. In the present study, LC–MS data indicated that abundant cyanidin-based anthocyanins accumulated in the petioles of purple celery (‘Nanxuan liuhe purple celery’). A gene encoding UDP-galactose: cyanidin 3-O-galactosyltransferase, namely AgUCGalT1, was isolated from purple celery and expressed in Escherichia coli BL21 (DE3). Sequence alignments revealed that the AgUCGalT1 protein contained a highly conserved putative secondary plant glycosyltransferase (PSPG) motif. The glycosylation product catalyzed by AgUCGalT1 was detected using UPLC equipment. The recombinant AgUCGalT1 had an optimal enzyme activity at 35 °C and pH 8.0, and showed highest enzyme activity toward cyanidin among the enzyme activities involving other substances, namely, peonidin, quercetin, and kaempferol. The expression levels of AgUCGalT1 were positively correlated with the total anthocyanin contents in purple and non-purple celery varieties. Crude enzymes extracted from purple celery exhibited glycosylation ability, whereas crude enzymes obtained from non-purple celery did not have this ability. This work provided evidence as a basis for investigations on the function of AgUCGalT1 in anthocyanin glycosylation in purple celery.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Abbreviations
- Fw:
-
Fresh weight
- Gln:
-
Glutamine
- His:
-
Histidine
- LC–MS:
-
Liquid chromatography–mass spectrometry
- ORF:
-
Open reading frame
- PSPG:
-
Putative secondary plant glycosyltransferase
- ROS:
-
Reactive oxygen species
- TF:
-
Transcription factor
- UDP:
-
Uridine diphosphate
- UGTs:
-
Uridine diphosphate glycosyltransferases
- UPLC:
-
Ultra-performance liquid chromatography
References
Ahmed NU, Park JI, Jung HJ, Hur Y, Nou IS (2015) Anthocyanin biosynthesis for cold and freezing stress tolerance and desirable color in Brassica rapa. Funct Integr Genom 15(4):383–394. https://doi.org/10.1007/s10142-014-0427-7
An JP, Qu FJ, Yao JF, Wang XN, You CX, Wang XF, Hao YJ (2017) The bZIP transcription factor MdHY5 regulates anthocyanin accumulation and nitrate assimilation in apple. Hortic Res 4:17023. https://doi.org/10.1038/hortres.2017.23
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254
Das SS, Gauri SS, Misra BB, Biswas M, Dey S (2013) Purification and characterization of a betanidin glucosyltransferase from Amaranthus tricolor L catalyzing non-specific biotransformation of flavonoids. Plant Sci 211:61–69. https://doi.org/10.1016/j.plantsci.2013.07.003
Dianat M, Veisi A, Ahangarpour A, Fathi Moghaddam H (2015) The effect of hydro-alcoholic celery (Apium graveolens) leaf extract on cardiovascular parameters and lipid profile in animal model of hypertension induced by fructose. Avicenna J Phytomed 5(3):203–209
Dougall DK, Baker DC, Gakh EG, Redus MA, Whittemore NA (1998) Studies on the stability and conformation of monoacylated anthocyanins part 2—anthocyanins from wild carrot suspension cultures acylated with supplied carboxylic acids. Carbohydr Res 310(3):177–189. https://doi.org/10.1016/S0008-6215(98)00124-4
Espley RV, Hellens RP, Putterill J, Stevenson DE, Kutty-Amma S, Allan AC (2007) Red colouration in apple fruit is due to the activity of the MYB transcription factor, MdMYB10. Plant J 49(3):414–427. https://doi.org/10.1111/j.1365-313X.2006.02964.x
Feng K, Xu ZS, Que F, Liu JX, Wang F, Xiong AS (2018) An R2R3-MYB transcription factor, OjMYB1, functions in anthocyanin biosynthesis in Oenanthe javanica. Planta 247(2):301–315. https://doi.org/10.1007/s00425-017-2783-8
Fukuchi-Mizutani M (2003) Biochemical and molecular characterization of a novel UDP-glucose:anthocyanin 3′-O-glucosyltransferase, a key enzyme for blue anthocyanin biosynthesis, from gentian. Plant Physiol 132(3):1652–1663. https://doi.org/10.1104/pp.102.018242
Glassgen WE, Seitz HU, Metzger JW (1992) High-performance liquid-chromatography electrospray mass-spectrometry and tandem mass-spectrometry of anthocyanins from plant-tissues and cell-cultures of Daucus Carota L. Biol Mass Spectrom 21(6):271–277. https://doi.org/10.1002/bms.1200210602
Gonzalez A, Zhao M, Leavitt JM, Lloyd AM (2008) Regulation of the anthocyanin biosynthetic pathway by the TTG1/bHLH/Myb transcriptional complex in Arabidopsis seedlings. Plant J 53(5):814–827. https://doi.org/10.1111/j.1365-313X.2007.03373.x
Harborne JB, Williams CA (2000) Advances in flavonoid research since 1992. Phytochemistry 55(6):481–504. https://doi.org/10.1016/S0031-9422(00)00235-1
He F, Mu L, Yan GL, Liang NN, Pan QH, Wang J, Reeves MJ, Duan CQ (2010) Biosynthesis of anthocyanins and their regulation in colored grapes. Molecules 15(12):9057–9091. https://doi.org/10.3390/molecules15129057
Hughes J, Hughes MA (1994) Multiple secondary plant product UDP-glucose glucosyltransferase genes expressed in cassava (Manihot esculenta Crantz) cotyledons. DNA Seq 5(1):41–49
Jaakola L, Maatta K, Pirttila AM, Torronen R, Karenlampi S, Hohtola A (2002) Expression of genes involved in anthocyanin biosynthesis in relation to anthocyanin, proanthocyanidin, and flavonol levels during bilberry fruit development. Plant Physiol 130(2):729–739. https://doi.org/10.1104/pp.006957
Jia XL, Wang GL, Xiong F, Yu XR, Xu ZS, Wang F, Xiong AS (2015) De novo assembly, transcriptome characterization, lignin accumulation, and anatomic characteristics: novel insights into lignin biosynthesis during celery leaf development. Sci Rep 5:8259. https://doi.org/10.1038/Srep08259
Jin W, Wang H, Li M, Wang J, Yang Y, Zhang X, Yan G, Zhang H, Liu J, Zhang K (2016) The R2R3 MYB transcription factor PavMYB10.1 involves in anthocyanin biosynthesis and determines fruit skin colour in sweet cherry (Prunus avium L.). Plant Biotechnol J 14(11):2120–2133. https://doi.org/10.1111/pbi.12568
Kobayashi S, Goto-Yamamoto N, Hirochika H (2004) Retrotransposon-induced mutations in grape skin color. Science 304(5673):982. https://doi.org/10.1126/science.1095011
Kogawa K, Kato N, Kazuma K, Noda N, Suzuki M (2007) Purification and characterization of UDP-glucose: anthocyanin 3′,5′-O-glucosyltransferase from Clitoria ternatea. Planta 226(6):1501–1509. https://doi.org/10.1007/s00425-007-0584-1
Kovinich N, Saleem A, Arnason JT, Miki B (2010) Functional characterization of a UDP-glucose:flavonoid 3-O-glucosyltransferase from the seed coat of black soybean (Glycine max (L.) Merr.). Phytochemistry 71(11–12):1253–1263. https://doi.org/10.1016/j.phytochem.2010.05.009
Kubo A, Arai Y, Nagashima S, Yoshikawa T (2004) Alteration of sugar donor specificities of plant glycosyltransferases by a single point mutation. Arch Biochem Biophys 429(2):198–203. https://doi.org/10.1016/j.abb.2004.06.021
Li YY, Mao K, Zhao C, Zhao XY, Zhang HL, Shu HR, Hao YJ (2012) MdCOP1 ubiquitin E3 ligases interact with MdMYB1 to regulate light-induced anthocyanin biosynthesis and red fruit coloration in apple. Plant Physiol 160(2):1011–1022. https://doi.org/10.1104/pp.112.199703
Li MY, Wang F, Jiang Q, Ma J, Xiong AS (2014) Identification of SSRs and differentially expressed genes in two cultivars of celery Apium graveolens L by deep transcriptome sequencing. Hortic Res 1:10. https://doi.org/10.1038/hortres.2014.10
Li MY, Wang F, Jiang Q, Wang GL, Tian C, Xiong AS (2016) Validation and comparison of reference genes for qPCR normalization of celery (Apium graveolens) at different development stages. Front Plant Sci 7:313. https://doi.org/10.3389/fpls.2016.00313
Li P, Li YJ, Zhang FJ, Zhang GZ, Jiang XY, Yu HM, Hou BK (2017) The Arabidopsis UDP-glycosyltransferases UGT79B2 and UGT79B3, contribute to cold, salt and drought stress tolerance via modulating anthocyanin accumulation. Plant J 89(1):85–103. https://doi.org/10.1111/tpj.13324
Li MY, Hou XL, Wang F, Tan GF, Xu ZS, Xiong AS (2018) Advances in the research of celery, an important Apiaceae vegetable crop. Crit Rev Biotechnol 38(2):172–183. https://doi.org/10.1080/07388551.2017.1312275
Lim EK, Ashford DA, Hou BK, Jackson RG, Bowles DJ (2004) Arabidopsis glycosyltransferases as biocatalysts in fermentation for regioselective synthesis of diverse quercetin glucosides. Biotechnol Bioeng 87(5):623–631. https://doi.org/10.1002/bit.20154
Matsui K, Umemura Y, Ohme-Takagi M (2008) AtMYBL2, a protein with a single MYB domain, acts as a negative regulator of anthocyanin biosynthesis in Arabidopsis. Plant J 55(6):954–967. https://doi.org/10.1111/j.1365-313X.2008.03565.x
Nakabayashi R, Yonekura-Sakakibara K, Urano K, Suzuki M, Yamada Y, Nishizawa T, Matsuda F, Kojima M, Sakakibara H, Shinozaki K, Michael AJ, Tohge T, Yamazaki M, Saito K (2014) Enhancement of oxidative and drought tolerance in Arabidopsis by overaccumulation of antioxidant flavonoids. Plant J 77(3):367–379. https://doi.org/10.1111/tpj.12388
Petroni K, Tonelli C (2011) Recent advances on the regulation of anthocyanin synthesis in reproductive organs. Plant Sci 181(3):219–229. https://doi.org/10.1016/j.plantsci.2011.05.009
Pourcel L, Routaboul JM, Cheynier V, Lepiniec L, Debeaujon I (2007) Flavonoid oxidation in plants: from biochemical properties to physiological functions. Trends Plant Sci 12(1):29–36. https://doi.org/10.1016/j.tplants.2006.11.006
Reed J (2002) Cranberry flavonoids, atherosclerosis and cardiovascular health. Crit Rev Food Sci Nutr 42(3 Suppl):301–316. https://doi.org/10.1080/10408390209351919
Rose A, Glassgen WE, Hopp W, Seitz HU (1996) Purification and characterization of glycosyltransferases involved in anthocyanin biosynthesis in cell-suspension cultures of Daucus carota L. Planta 198(3):397–403
Schmittgen TD, Livak KJ (2008) Analyzing real-time PCR data by the comparative C(T) method. Nat Protoc 3(6):1101–1108
Schwede T, Kopp J, Guex N, Peitsch MC (2003) SWISS-MODEL: an automated protein homology-modeling server. Nucleic Acids Res 31(13):3381–3385
Shi MZ, Xie DY (2014) Biosynthesis and metabolic engineering of anthocyanins in Arabidopsis thaliana. Recent Pat Biotechnol 8(1):47–60
Shih PH, Chan YC, Liao JW, Wang MF, Yen GC (2010) Antioxidant and cognitive promotion effects of anthocyanin-rich mulberry (Morus atropurpurea L.) on senescence-accelerated mice and prevention of Alzheimer’s disease. J Nutr Biochem 21(7):598–605. https://doi.org/10.1016/j.jnutbio.2009.03.008
Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28(10):2731–2739. https://doi.org/10.1093/molbev/msr121
Tan GF, Ma J, Zhang XY, Xu ZS, Xiong AS (2017a) AgFNS overexpression increase apigenin and decrease anthocyanins in petioles of transgenic celery. Plant Sci 263:31–38. https://doi.org/10.1016/j.plantsci.2017.07.001
Tan GF, Wang F, Ma J, Zhang XY, Xiong AS (2017b) Analysis of anthocyanin and apigenin contents and the expression profiles of biosynthesis-related genes in the purple and non-purple varieties of celery. Acta Hortic Sin 44(7):1327–1334. https://doi.org/10.16420/j.issn.0513-353x.2017-0221
Tanaka Y, Sasaki N, Ohmiya A (2008) Biosynthesis of plant pigments: anthocyanins, betalains and carotenoids. Plant J 54(4):733–749. https://doi.org/10.1111/j.1365-313X.2008.03447.x
Vogt T, Jones P (2000) Glycosyltransferases in plant natural product synthesis: characterization of a supergene family. Trends Plant Sci 5(9):380–386
Walker AR, Davison PA, Bolognesi-Winfield AC, James CM, Srinivasan N, Blundell TL, Esch JJ, Marks MD, Gray JC (1999) The TRANSPARENT TESTA GLABRA1 locus, which regulates trichome differentiation and anthocyanin biosynthesis in Arabidopsis, encodes a WD40 repeat protein. Plant Cell 11(7):1337–1350
Winkel-Shirley B (2001) Flavonoid biosynthesis. A colorful model for genetics, biochemistry, cell biology, and biotechnology. Plant Physiol 126(2):485–493
Xie DY, Sharma SB, Dixon RA (2004) Anthocyanidin reductases from Medicago truncatula and Arabidopsis thaliana. Arch Biochem Biophys 422(1):91–102. https://doi.org/10.1016/j.abb.2003.12.011
Xu ZS, Xue W, Xiong AS, Lin YQ, Xu J, Zhu B, Zhao W, Peng RH, Yao QH (2013) Characterization of a bifunctional O- and N-glucosyltransferase from Vitis vinifera in glucosylating phenolic compounds and 3,4-dichloroaniline in Pichia pastoris and Arabidopsis thaliana. PLoS One 8(11):e80449. https://doi.org/10.1371/journal.pone.0080449
Xu ZS, Huang Y, Wang F, Song X, Wang GL, Xiong AS (2014) Transcript profiling of structural genes involved in cyanidin-based anthocyanin biosynthesis between purple and non-purple carrot (Daucus carota L.) cultivars reveals distinct patterns. BMC Plant Biol 14:262. https://doi.org/10.1186/S12870-014-0262-Y
Xu ZS, Ma J, Wang F, Ma HY, Wang QX, Xiong AS (2016) Identification and characterization of DcUCGalT1, a galactosyltransferase responsible for anthocyanin galactosylation in purple carrot Daucus carota L taproots. Sci Rep 6:27356. https://doi.org/10.1038/srep27356
Zhou J, Miners JO (2014) Enzyme kinetics of uridine diphosphate glucuronosyltransferases (UGTs). Methods Mol Biol 1113:203–228. https://doi.org/10.1007/978-1-62703-758-7_11
Acknowledgements
The research was supported by the New Century Excellent Talents in University (NCET-11-0670); National Natural Science Foundation of China (31272175; 31501775); Jiangsu Natural Science Foundation (BK20130027); Priority Academic Program Development of Jiangsu Higher Education Institutions.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Feng, K., Xu, ZS., Liu, JX. et al. Isolation, purification, and characterization of AgUCGalT1, a galactosyltransferase involved in anthocyanin galactosylation in purple celery (Apium graveolens L.). Planta 247, 1363–1375 (2018). https://doi.org/10.1007/s00425-018-2870-5
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00425-018-2870-5