Cashew-nut tree exudate gum: Identification of an arabinogalactan-protein as a constituent of the gum and use on the stimulation of somatic embryogenesis

doi:10.1016/j.plantsci.2007.07.008

Plant Science

Volume 173, Issue 4, October 2007, Pages 468-477

https://doi.org/10.1016/j.plantsci.2007.07.008 Get rights and content

Abstract

Arabinogalactan-proteins (AGPs) are a class of plant proteoglycans. In this work, we show that gel permeation chromatography of the cashew-nut tree exudate gum (CNTG), previously characterized as an acidic arabinogalactan (type II), yielded a fraction named fraction III. SDS-PAGE analysis of CNTG has an AGP-like smear, weakly stained by the β-glucosyl Yariv reagent. CNTG and fraction III reacted strongly with the JIM13 anti-AGP monoclonal antibody and presented a protein content of 1.0 and 0.2% (w/w), respectively. CNTG and fraction III showed a high Hyp content (∼50%), and Ser, Pro, Thr and Leu as the most abundant amino acids. Analysis of the carbohydrate structure, its extracellular location and low protein content, and the amino acid composition indicate that CNTG contains at least one fraction that fits the diagnostic features of a “classical” AGP. CNTG-enriched (0.1 μg ml⁻¹) culture medium stimulated carrot somatic embryogenesis after 2 and 3 weeks of culture. The conversion of somatic embryos into plantlets was also enhanced in CNTG-enriched culture media, making this gum a potential tool for the improvement of conversion rates in systems in which low conversion rates are a constraint to micropropagation and breeding programs.

Introduction

The carbohydrate moiety of the exudate gum from cashew-nut trees (Anacardium occidentale L.) grown in Papua New Guinea [1], India [1], [2], Venezuela [3] and Brazil [4], have been extensively studied from both composition and structural standpoint. The structural analysis of the major fraction present in the Brazilian cashew-nut tree exudate gum (CNTG) revealed that it is an type II arabinogalactan with a (1 → 3)-linked β-d-Galp main chain, highly substituted at O-6 by various side-chains [4]. Although the protein content of CNTG from cashew-nut trees from India and Papua has been reported (1–3%, w/v) [1], no information about the chemical amino acid composition of the protein moiety of this gum exudate has been reported.

The cashew-nut tree gum exudate (CNTG) has been previously demonstrated to display potential rheological applications [5] and crop protection against insect [6]. In addition, properties against virus [7], bacteria and fungi [6] for CNTG have also been demonstrated.

In this paper, we describe the identification of a classical AGP as a constituent of the CNTG. In addition, we corroborate earlier reports demonstrating a biotechnological use for AGPs, the improvement of somatic embryo formation. However, more importantly, we describe, for the first time, an AGP-driven enhancement of the conversion rate of somatic embryos into plantlets.

Section snippets

Fractionation of CNTG by gel permeation chromatography

The cashew-nut tree gum exudate (CNTG) was harvested at Caruaru County (State of Pernambuco, Brazil) by hand during the summer of 2001. CNTG was precipitated in ethanol (4 vol). Afterwards, the precipitate (0.5 g) was dissolved in 8 M guanidine. HCl and subjected to gel permeation chromatography (GPC, 20 mm × 800 mm, Toyopearl HW-65F, TosoHaas, Stuttgart, Germany) using MilliQ water as the eluent. Each GPC fraction (5 ml) was monitored at A_215 nm and A_280 nm, for protein detection. In addition to the

Fractionation of CNTG by gel permeation chromatography (GPC)

The GPC fractionation of CNTG showed a single major peak for the protein profile (Fig. 1), whereas the fractions eluting from 100 to 240 ml all tested positive for carbohydrate. On the basis of the chromatographic profile (i.e. protein and total carbohydrate content) obtained the eluted fractions were pooled to generate four fractions, I, II, III and IV (see Fig. 1). After desalting, the yield for fractions I, II, III and IV (compared to CNTG) was 12%, 21%, 66% and 1%, respectively. Since

Identification of an arabinogalactan-protein as a constituent of CNTG

d-Galactose and l-arabinose are the major monosaccharides found in type II AG and AGPs, with most samples containing more Gal than Ara [19]. The structure of the carbohydrate moiety of the cashew-nut tree exudate gum (CNTG) has been previously described [4]. The structural analysis of the CNTG revealed that it has a (1 → 3)-linked β-d-Galp main chain, highly substituted at O-6 by Gal, Ara, Glc, Rha and GlcA residues [4]. When taken together, these data suggest CNTG belongs to the type II class of

Acknowledgements

The authors thank CNPq-Brazil and CAPES-Brazil for financial support. We also thank Ms. Naomi Ling and Dr. Shaio-Lim Mau (CRC for Bioproducts, School of Botany, The University of Melbourne, Australia), for the gift of dGA and help with the RP-HPLC analysis, respectively. Dr. Tracy Putoczki (PCBRC, The University of Melbourne, Australia) is also acknowledged for assistance with the immuno-dot blotting assay. FP and AB acknowledge the support of the CRC for Bioproducts.

References (39)

D.M.W. Anderson et al.
Uronic acid materials XLVI. Composition of gum exudate from Anacardium occidentale
Phytochemistry
(1974)
D.M.W. Anderson et al.
Uronic acid materials XLVIII. Structural analysis of the gum polysaccharides from Anacardium occidentate
Anal. Chim. Acta
(1975)
G.L. De Pinto et al.
Comparison of three Anacardiaceae gum exudates
Biochem. System. Ecol.
(1995)
J.M. Menestrina et al.
Similarity of monosaccharide, oligosaccharide and polysaccharide structures in gum exudate of Anacardium occidentale
Phytochemistry
(1998)
M.B. Zakaria et al.
Rheological properties of cashew
Carbohydr. Polym.
(1996)
J.L.S. Gonçalves et al.
In vitro anti-rotavirus activity of some medicinal plants used in Brazil against diarrhea
J. Ethnopharmacol.
(2005)
G. Van-Holst et al.
Quantification of arabinogalactan-protein in plant extracts by single radial gel diffusion
Anal. Biochem.
(1985)
M.M. Bradford
A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding
Anal Biochem.
(1976)
J. Defaye et al.
Structural studies of gum arabic, the exudates polysaccharide from Acacia senegal
Carbohydr. Res.
(1986)
R.L. Whistler
Exudate gums

L.J. Goodrum et al.

Gum arabic glycoprotein contains glycomodules of both extensin and arabinogalactan-glycoproteins

Phytochemistry

(2000)

S. Thude et al.

High molecular weight constituents from roots of Echinacea pallida: an arabinogalactan-protein and an arabinan

Phytochemistry

(2005)

P. Komalavilas et al.

Arabinogalactan-proteins from the suspension culture medium and plasma membrane of rose cells

J. Biol. Chem.

(1991)

Y. Akiyama et al.

An extracellular arabinogalactan-protein from Nicotiana tabacum

Phytochemistry

(1981)

M.R. Marques et al.

Antimicrobial and insecticidal activities of cashew tree gum exudates

Ann. Appl. Biol.

(1992)

M. Dubois et al.

Colorimetric method for the determination of sugars and related substances

Anal. Chem.

(1956)

T.M.C.C. Filizetti-Cozzi et al.

Measurement of uronic. acids without interference from neutral sugars

Anal. Biochem.

(1991)

J. Yariv et al.

The interaction of glycosides and saccharides with antibody to the corresponding phenylazo glycosides

Biochem. J.

(1962)

J.P. Knox et al.

Developmentally regulated epitopes of the cell surface arabinogalactan proteins and their relation to root tissue pattern formation

Plant J.

(1991)

Cited by (24)

Enzymatic synthesis of prebiotic galactooligosaccharides from galactose derived from gum arabic
2023, Food Chemistry
A novel enzymatic process was established for galactooligosaccharides (GOS) synthesis by using plant-derived galactose as substrate, without producing any byproducts. The galactose was prepared from the acid hydrolysate of gum arabic. The yeast Kluyveromyces lactis producing β-galactosidase capable of catalyzing GOS synthesis from galactose was screened out. The synthesis conditions using the yeast cells as enzyme source were optimized by both single-factor experiment and response surface methodology, with the highest GOS yield reached 45%. The composition of reaction mixture contained only GOS and unreacted galactose, which could be easily separated by the cation exchange resin column. The structures of major GOS products were identified as Gal-β-D-(1 → 6)-Gal, Gal-β-D-(1 → 3)-Gal, and Gal-β-D-(1 → 6)-Gal-β-D-(1 → 6)-Gal by MS and NMR spectra. Moreover, the β-galactosidase-containing cells can be recycled for at least 30 batches of GOS synthesis at 35 °C, with the enzyme activity remaining above 60%.
Plants arabinogalactans: From structures to physico-chemical and biological properties
2021, Biotechnology Advances
Arabinogalactans (AGs) are plant heteropolysaccharides with complex structures occasionally attached to proteins (AGPs). AGs in cell matrix of different parts of plant are freely available or chemically bound to pectin rhamnogalactan. Type I with predominantly β-d-(1 → 4)-galactan and type II with β-d-(1 → 3) and/or (1 → 6)-galactan structural backbones construct the two main groups of AGs. In the current review, the chemical structure of AGs is firstly discussed focusing on non-traditional plant sources and not including well known industrial gums. After that, processes for their extraction and purification are considered and finally their techno-functional and biological properties are highlighted. The role of AG structure and function on health advantages such as anti-tumor, antioxidant, anti-ulcer- anti-diabetic and other activites and also the immunomodulatory effects on in-vivo model systems are overviewed.
β-1,3-Glucanases and chitinases participate in the stress-related defence mechanisms that are possibly connected with modulation of arabinogalactan proteins (AGP) required for the androgenesis initiation in rye (Secale cereale L.)
2021, Plant Science
This work presents the biochemical, cytochemical and molecular studies on two groups of PR proteins, β-1,3-glucanases and chitinases, and the arabinogalactan proteins (AGP) during the early stages of androgenesis induction in two breeding lines of rye (Secale cereale L.) with different androgenic potential. The process of androgenesis was initiated by tillers pre-treatments with low temperature, mannitol and/or reduced glutathione and resulted in microspores reprogramming and formation of androgenic structures what was associated with high activity of β-1,3-glucanases and chitinases. Some isoforms of β-1,3-glucanases, namely several acidic isoforms of about 26 kDa; appeared to be anther specific. Chitinases were well represented but were less variable. RT-qPCR revealed that the cold-responsive chitinase genes Chit1 and Chit2 were expressed at a lower level in the microspores and whole anthers while the cold-responsive Glu2 and Glu3 were not active. The stress pre-treatments modifications promoted the AGP accumulation. An apparent dominance of some AGP epitopes (LM2, JIM4 and JIM14) was detected in the androgenesis-responsive rye line. An abundant JIM13 epitopes in the vesicles and inner cell walls of the microspores and in the cell walls of the anther cell layers appeared to be the most specific for embryogenesis.
Anti-proliferative profile of Anacardium occidentale polysaccharide and characterization by AFM
2020, International Journal of Biological Macromolecules
This paper explores the application of cashew gum (CG) as an in vitro antiproliferative, firstly by isolating and characterizing the gum using elemental analysis, gel-permeation chromatography, nuclear magnetic resonance (NMR) and atomic force microscopy (AFM). The molar mass of isolated CG was in the order of 10³–10⁴ g/mol, with small protein traces present. Polymer characterization by NMR identified key signals correlating to galactose, glucose, rhamnose and acid-related groups. Three distinct conformational stages were observed by AFM. The impact of CG on cell morphology and viability with both tumor and non-tumor cell lines was studied by AFM and 3-(4,5-dimethyl-2-thiazole)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay respectively. Antiproliferative activity was confirmed for HCT116 (colorectal carcinoma), B16F10 (melanoma) and HL60 (promyelocytic leukemia) cancer cell lines. A change in cell morphology was demonstrated as an increased surface roughness for HL60. Considering that a CG does not exhibit cytotoxicity to non-tumor lines, it can be seen that the CG shows selectivity for tumor cells and can be a promising biomaterial for future studies.
Structural features of a new water-soluble polysaccharide from the gum exudates of Amygdalus scoparia Spach (Zedo gum)
2018, Carbohydrate Polymers
Citation Excerpt :
This information confirmed the result from FT-IR analysis regarding the presence of pyranose and furanose forms sugars in CZGS-1. In the 13C NMR spectrum of CZGS-1 (Fig. 4B), the resonances due to C-1 of β-d-Galp residues were observed at 104.1, 104.3 and 104.6 ppm (Cui, 2005; Pereira-Netto et al., 2007; Sims & Furneaux, 2003; Yuan et al., 2016). The other anomeric carbon signals at 103.6, 104.2 and 101.5 ppm were assigned to anomeric carbons of β-d-Xylp (Fischer et al., 2004; Yin et al., 2012), β-d-GlcAp and α-l-Rhap residues (Adinolfi et al., 1994; Delgobo, Gorin, Tischer, & Iacomini, 1999; Pereira-Netto et al., 2007; Sims & Furneaux, 2003), respectively.
A new water-soluble polysaccharide (CZGS-1) with molecular weight of 4860 kDa and a specific optical rotation of +31.5° (c 1.0, H₂O), was extracted from the gum exudate of Amygdalus scoparia Spach by hot water, following by purification with DEAE-cellulose A52 and Sephacryl S-400 HR columns. Monosaccharide composition analysis indicates that CZGS-1 was an arabino galactan including Rha, Ara, Xyl, and Gal with a relative molar ratio of 1.1:20:5.2:17.9. The uronic acid content of CZGS-1 was about 6%. Structural features of CZGS-1 was investigated by a combination of partial acid hydrolysis, methylation and GC–MS analysis, periodic acid oxidation and Smith degradation, FT-IR and NMR spectroscopy. The results indicated that CZGS-1 possesses a backbone of →3,6)-β-d-Galp-(1→, →3)-β-d-Galp-(1→, and →3)-α-l-Araf-(1→ residues with side chains attached to O-3 and O-6 positions of 1,3,6-linked β-d-Galp. The side chains are consisted of β-d-Xylp-(1 → 3)-α-l-Araf-(1 → 3)-α-l-Araf-(1→), α-l-Rhap-(1 → 6)-β-d-Galp-(1→), and β-d-GlcAp-(1 → 6)-β-d-Galp-(1→).
Development and characterization of bacterial cellulose produced by cashew tree residues as alternative carbon source
2017, Industrial Crops and Products
Bacterial cellulose (BC) has been extensively exploited for applications in materials science, biomedical and technological fields. The BC production demands culture media rich in carbon sources. Agro-forestry residues constitute an interesting source of nutrients for microorganism, but they are frequently wasted. For cashew crop, exudate is periodically extracted from the tree trunks to increase the production of cashew nut, the most valuable product from cashew trees that produces about 700 g of exudate/year, which remains wasted. Here, we associated the nutritional properties of residues from cashew tree with the need of carbon sources for BC, in attempt to valorize the residue and to decrease the costs of BC production. The carbon source from Hestrin Schramm culture medium was totally or partially replaced by cashew tree residues and the BC production was evaluated. The produced BC membrane in static medium was characterized by FTIR, SEM and TGA and the kinetics of production was determined, suggesting the cashew tree residues as a potential carbon source for BC production.

View all citing articles on Scopus

View full text

Cashew-nut tree exudate gum: Identification of an arabinogalactan-protein as a constituent of the gum and use on the stimulation of somatic embryogenesis

Abstract

Introduction

Section snippets

Fractionation of CNTG by gel permeation chromatography

Fractionation of CNTG by gel permeation chromatography (GPC)

Identification of an arabinogalactan-protein as a constituent of CNTG

Acknowledgements

Phytochemistry

Anal. Chim. Acta

Biochem. System. Ecol.

Phytochemistry

Carbohydr. Polym.

J. Ethnopharmacol.

Anal. Biochem.

Anal Biochem.

Carbohydr. Res.

Phytochemistry

Phytochemistry

J. Biol. Chem.

Phytochemistry

Antimicrobial and insecticidal activities of cashew tree gum exudates

Ann. Appl. Biol.

Colorimetric method for the determination of sugars and related substances

Anal. Chem.

Measurement of uronic. acids without interference from neutral sugars

Anal. Biochem.

The interaction of glycosides and saccharides with antibody to the corresponding phenylazo glycosides

Biochem. J.

Developmentally regulated epitopes of the cell surface arabinogalactan proteins and their relation to root tissue pattern formation

Plant J.