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Arabinosylation Plays a Crucial Role in Extensin Cross-linking In Vitro.

Chen Y, Dong W, Tan L, Held MA, Kieliszewski MJ - Biochem Insights (2015)

Bottom Line: Proposed functions of EXT arabinosylation include stabilizing the polyproline II helix structure and facilitating EXT cross-linking.Our results indicate that EXT arabinosylation is required for EXT cross-linking in vitro and the fourth arabinosyl residue in the tetraarabinoside chain, which is uniquely α-linked, may determine the initial cross-linking rate.Our results also confirm the conserved structure of the oligoarabinosides across species, indicating an evolutionary significance for EXT arabinosylation.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, Ohio University, Athens, OH, USA.

ABSTRACT
Extensins (EXTs) are hydroxyproline-rich glycoproteins (HRGPs) that are structural components of the plant primary cell wall. They are basic proteins and are highly glycosylated with carbohydrate accounting for >50% of their dry weight. Carbohydrate occurs as monogalactosyl serine and arabinosyl hydroxyproline, with arabinosides ranging in size from ~1 to 4 or 5 residues. Proposed functions of EXT arabinosylation include stabilizing the polyproline II helix structure and facilitating EXT cross-linking. Here, the involvement of arabinosylation in EXT cross-linking was investigated by assaying the initial cross-linking rate and degree of cross-linking of partially or fully de-arabinosylated EXTs using an in vitro cross-linking assay followed by gel permeation chromatography. Our results indicate that EXT arabinosylation is required for EXT cross-linking in vitro and the fourth arabinosyl residue in the tetraarabinoside chain, which is uniquely α-linked, may determine the initial cross-linking rate. Our results also confirm the conserved structure of the oligoarabinosides across species, indicating an evolutionary significance for EXT arabinosylation.

No MeSH data available.


Characterized primary structure of RSH Hyp-Ara4 and Hyp-Ara3. Each arabinose residue is labeled corresponding to those in Table 5 with their anomeric configurations. The glycosidic linkages between arabinose residues and with Hyp are labeled according to the HMBC data (Figs. 4 and 5) with carbon numbers indicating the C atoms involved in the linkages.
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f6-bci-suppl.2-2015-001: Characterized primary structure of RSH Hyp-Ara4 and Hyp-Ara3. Each arabinose residue is labeled corresponding to those in Table 5 with their anomeric configurations. The glycosidic linkages between arabinose residues and with Hyp are labeled according to the HMBC data (Figs. 4 and 5) with carbon numbers indicating the C atoms involved in the linkages.

Mentions: TFA elution of RSH Hyp-oligoarabinosides yielded a profile similar to that of HCl elution (Fig. 1A, first panel). Peaks containing Hyp-Ara4 and Hyp-Ara3 were collected (cis and trans peaks pooled for NMR analysis), and through NMR spectra (COSY, TCOSY, HSQC, and HMBC) the glycosyl residues, ring systems, and ordered linkages of the components of RSH Hyp-Ara4 (Fig. 4) and Hyp-Ara3 (Fig. 5) were identified. The results corroborate the chemical shift obtained earlier by Akiyama et al50,51 for Hyparabinosides isolated from tobacco suspension culture cell walls (Table 5) and identified the structures of RSH Hyp-Ara4 and Hyp-Ara3 as (Fig. 6).


Arabinosylation Plays a Crucial Role in Extensin Cross-linking In Vitro.

Chen Y, Dong W, Tan L, Held MA, Kieliszewski MJ - Biochem Insights (2015)

Characterized primary structure of RSH Hyp-Ara4 and Hyp-Ara3. Each arabinose residue is labeled corresponding to those in Table 5 with their anomeric configurations. The glycosidic linkages between arabinose residues and with Hyp are labeled according to the HMBC data (Figs. 4 and 5) with carbon numbers indicating the C atoms involved in the linkages.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4629521&req=5

f6-bci-suppl.2-2015-001: Characterized primary structure of RSH Hyp-Ara4 and Hyp-Ara3. Each arabinose residue is labeled corresponding to those in Table 5 with their anomeric configurations. The glycosidic linkages between arabinose residues and with Hyp are labeled according to the HMBC data (Figs. 4 and 5) with carbon numbers indicating the C atoms involved in the linkages.
Mentions: TFA elution of RSH Hyp-oligoarabinosides yielded a profile similar to that of HCl elution (Fig. 1A, first panel). Peaks containing Hyp-Ara4 and Hyp-Ara3 were collected (cis and trans peaks pooled for NMR analysis), and through NMR spectra (COSY, TCOSY, HSQC, and HMBC) the glycosyl residues, ring systems, and ordered linkages of the components of RSH Hyp-Ara4 (Fig. 4) and Hyp-Ara3 (Fig. 5) were identified. The results corroborate the chemical shift obtained earlier by Akiyama et al50,51 for Hyparabinosides isolated from tobacco suspension culture cell walls (Table 5) and identified the structures of RSH Hyp-Ara4 and Hyp-Ara3 as (Fig. 6).

Bottom Line: Proposed functions of EXT arabinosylation include stabilizing the polyproline II helix structure and facilitating EXT cross-linking.Our results indicate that EXT arabinosylation is required for EXT cross-linking in vitro and the fourth arabinosyl residue in the tetraarabinoside chain, which is uniquely α-linked, may determine the initial cross-linking rate.Our results also confirm the conserved structure of the oligoarabinosides across species, indicating an evolutionary significance for EXT arabinosylation.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, Ohio University, Athens, OH, USA.

ABSTRACT
Extensins (EXTs) are hydroxyproline-rich glycoproteins (HRGPs) that are structural components of the plant primary cell wall. They are basic proteins and are highly glycosylated with carbohydrate accounting for >50% of their dry weight. Carbohydrate occurs as monogalactosyl serine and arabinosyl hydroxyproline, with arabinosides ranging in size from ~1 to 4 or 5 residues. Proposed functions of EXT arabinosylation include stabilizing the polyproline II helix structure and facilitating EXT cross-linking. Here, the involvement of arabinosylation in EXT cross-linking was investigated by assaying the initial cross-linking rate and degree of cross-linking of partially or fully de-arabinosylated EXTs using an in vitro cross-linking assay followed by gel permeation chromatography. Our results indicate that EXT arabinosylation is required for EXT cross-linking in vitro and the fourth arabinosyl residue in the tetraarabinoside chain, which is uniquely α-linked, may determine the initial cross-linking rate. Our results also confirm the conserved structure of the oligoarabinosides across species, indicating an evolutionary significance for EXT arabinosylation.

No MeSH data available.