Two Chains of Rhamnogalacturonan II Are Cross-Linked by Borate-Diol Ester Bonds in Higher Plant Cell Walls

Abstract

Polysaccharide moiety of the boron-polysaccharide complex (T. shown to be rhamnogalacturonan II by glycosyl-linkage analysis and the presence of diagnostic monosaccharides, including apiose, aceric acid, 2-Omethylfucose, and 3-deoxy-~-mann~2-octulosonic acid. Remova1 of boron from the complex reduced the molecular weight by one-half without causing a significant increase in the number of reducing end groups, indicating that boron, as boric acid, links two rhamnogaladuronan II chains together to form the boron-polysaccharide complex.Boron (B) is an essential microelement for higher plants (reviewed by Loomis and Durst, 1992); however, its primary function is not known. A number of metabolic disorders that are consequences of B deficiency have been discussed in recent reviews (Dugger, 1983;Parr and Loughman, 1983; Loomis and Durst, 1992). Skok and McIlrath (1958) pointed out that the locus of B in the plant cell was little known, and the issue has never been resolved. We have shown that in cultured tobacco cells more than 98% of the B is present in the cell wall (Matoh et al., 1992), and that a BPC can be isolated from radish (Xapkanus sativus) cell walls (Matoh et al., 1993a). In this paper, the structure of the polysaccharide moiety of the radish BPC is reported and its physiological function is discussed. MATERIALS AND METHODS Preparation of the BPCThe BPC was prepared from radish (Xaphanus sativus) roots as described previously (Matoh et al., 1993a) with some modifications. Briefly, radish roots were grated and squeezed, and the macerated tissue was freezedried. The dried tissue (4 g) was treated with Pectinase-SS (0.lY0, w/v; Kyowa Chemical Products, NishiNakajima, Osaka, Japan) in 400 mL of 20 mM sodium acetate, pH 4.0, for 48 h at 25°C on a rotary shaker (130 rpm). The suspension was centrifuged min), and the supernatant was adjusted to pH 8.0 with 2 M Tris and then applied to a DEAE-Sepharose column (4.8 X 60 cm, C1-form, Pharmacia) equilibrated with 20 mM Tris-HC1, pH 8.0. The column was eluted with a 6-L linear gradient of O to 0.5 M NaCl in the column buffer, and the fractions containing B were pooled and dialyzed. The BPC was purified by rechromatography on the same DEAE-Sepharose column. Fractions containing B were subjected to gel filtration using a Superdex 75 column (2.6 X 60 cm, Pharmacia) equilibrated with 20 mM Tris-HC1, pH 8.0, containing 0.1 M NaC1. B-rich fractions were pooled, dialyzed against water, and lyophilized. Partia1 Hydrolysis of the BPC with AcidThe BPC (1 mg) was incubated in 0.1 M HCl(1 mL) for 15 min at 25OC. After neutralization with NaOH, an aliquot (100 pL) was analyzed by size-exclusion chromatography (YMC-pack Diol-120, 300 X 8 mm [YMC, Karasuma-Oike, Kyoto, Japan], Shimadzu 6A HPLC system). The column was equilibrated and eluted at a flow rate of 0.5 mL min-' with 50 mM sodium acetate, pH 5.2, containing 0.2 M NaC1. Saccharides were detected fluorometrically (Shimadzu RF 530 detector) after postcolumn labeling of their reducing termini with 2-cyanoacetamide (Honda et al., ...

Occurrence of the Primary Cell Wall Polysaccharide Rhamnogalacturonan II in Pteridophytes, Lycophytes, and Bryophytes. Implications for the Evolution of Vascular Plants

“…To confirm that the pteridophyte RG-II exists as a borate, cross-linked dimer solutions of the EPG-soluble material from P. nudum and E. hyemale were treated for 45 min at 22°C with 0.1 m HCl. These conditions, which are known to hydrolyze the borate diester and generate monomeric RG-II and boric acid (Ishii and Matsunaga, 1996;Kobayashi et al, 1996;O'Neill et al, 1996), convert the pteridophyte RG-II dimer into monomer (Fig. 4, A and B, middle profiles).…”

“…The 11 B-NMR spectrum of C. thalictroides RG-II (Fig. 4C) contains a signal at ␦-9.6 that originates from a 1:2 borate:diol diester (Ishii and Matsunaga, 1996;Kobayashi et al, 1996;O'Neill et al, 1996). To confirm that the pteridophyte RG-II exists as a borate, cross-linked dimer solutions of the EPG-soluble material from P. nudum and E. hyemale were treated for 45 min at 22°C with 0.1 m HCl.…”

“…The primary walls of seed-bearing tracheophytes (gymnosperms and angiosperms) are composed predominantly of cellulose, hemicellulose, and pectin together with lesser amounts of structural glycoproteins, minerals, enzymes, and phenolic esters (Carpita and Gibeaut, 1993 Recent studies have shown that boron has a major role in maintaining wall structure and that the boron requirements of flowering plants are correlated with the amounts of pectin in their primary walls (Loomis and Durst, 1992;Hu et al, 1996;Brown et al, 2002). Boron exists in primary walls predominantly as a 1:2 borate-diol ester that covalently cross-links two chains of the pectic polysaccharide rhamnogalacturonan II (RG-II; Kobayashi et al, 1996;Ishii and Matsunaga, 1996;O'Neill et al, 1996), which has the added effect of covalently cross-linking the two homogalacturonan chains upon which the RG-II molecules were constructed .…”

Functional Characterization of UDP-apiose Synthases from Bryophytes and Green Algae Provides Insight into the Appearance of Apiose-containing Glycans during Plant Evolution

“…Apiose links two side chains (A and B) to the galacturonan backbone of RG-II. The apiosyl residues of side chain A in two RG-II molecules are cross-linked by a borate diester to form the RG-II dimer (7)(8)(9)(10). At least 90% of the RG-II in primary walls exists as a dimer (11), and a reduction in the extent of RG-II cross-linking typically results in the formation of abnormal cell walls (12).…”

Distribuição de nutrientes e sintomas visuais de deficiência de boro em raízes de coqueiro-anão verde

“…Nas raízes grossas, mais lignificadas, parte do nutriente pode ter se acumulado nas paredes celulares, ligando-se com a pectina (Kobayashi et al,1996;Ishii & Matsunaga, 1996;Fleischer et al, 1998), e uma certa quantidade de B estaria livre para ser translocada. Dannel et al (2000) encontraram resultados mostrando que o B ligado à parede celular de plantas de girassol não é totalmente imóvel, uma vez que aproximadamente 28 % desse nutriente inicialmente presente na parede celular foi trocado ou liberado após três horas de omissão do B da solução nutritiva, ficando o restante aderido.…”