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Characterization of the hydroxyproline-rich protein core of an arabinogalactan-protein secreted from suspension-cultured Lolium multiflorum (Italian ryegrass) endosperm cells.

ABSTRACT: An arabinogalactan-protein (AGP) purified from the filtrate of liquid-suspension-cultured Italian-ryegrass (Lolium multiflorum) endosperm cells by affinity chromatography on myeloma protein J539-Sepharose was deglycosylated with trifluoromethanesulphonic acid to remove polysaccharide chains that are covalently associated with hydroxyproline residues in the peptide component of the proteoglycan. The protein core, which accounts for less than 10% (w/w) of the intact proteoglycan, was purified by h.p.l.c. It has an apparent Mr of 35,000, but reacts very poorly with both Coomassie Brilliant Blue R and silver stains. Amino-acid-sequence analysis of the N-terminus of the h.p.l.c.-purified protein core and of tryptic peptides generated from the unpurified protein reveals a high content of hydroxyproline and alanine. These are sometimes arranged in short (Ala-Hyp) repeat sequences of up to six residues. Polyclonal antibodies raised against the protein core do not cross-react with native AGP, the synthetic peptide (Ala-Hyp)4, poly-L-hydroxyproline or poly-L-proline. The results suggest that the polysaccharide chains in the native AGP render the protein core of the proteoglycan inaccessible to the antibodies and that the immunodominant epitopes include domains of the protein other than those rich in Ala-Hyp repeating units.

SUBMITTER: Gleeson PA

PROVIDER: S-EPMC1133664 | biostudies-other | 1989 Dec

REPOSITORIES: biostudies-other

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Biosynthesis of arabinogalactan-protein in Lolium multiflorum (Italian ryegrass) endosperm cells. Subcellular distribution of galactosyltransferases.

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Project description:AimMagnesium (Mg) deficiency (known as grass tetany) is a serious metabolic disorder that affects grazing ruminants. We tested whether Mg-fertiliser can increase Mg concentration of Italian ryegrasses (Lolium multiflorum L.) including a cultivar (cv. Bb2067; 'Magnet'), bred to accumulate larger concentrations of Mg.MethodsUnder controlled environment (CE) conditions, three cultivars (cv. Bb2067, cv. Bb2068, cv. RvP) were grown in low-nutrient compost at six fertiliser rates (0-1500 μM MgCl2.6H2O). Under field conditions, the three cultivars in the CE condition and cv. Alamo were grown at two sites, and four rates of MgSO4 fertiliser application rates (0-200 kg ha-1 MgO). Multiple grass cuts were taken over two-years.ResultsGrass Mg concentration increased with increasing Mg-fertiliser application rates in all cultivars and conditions. Under field conditions, cv. Bb2067 had 11-73% greater grass Mg concentration and smaller forage tetany index (FTI) than other cultivars across the Mg-fertiliser application rates, sites and cuts. Grass dry matter (DM) yield of cv. Bb2067 was significantly (p < 0.05) smaller than cv. Alamo. The effect of Mg-fertiliser rate on DM yield was not significant (p ≥ 0.05).ConclusionsBiofortification of grass with Mg through breeding and agronomy can improve the forage Mg concentration for grazing ruminants, even in high-growth spring grass conditions when hypomagnesaemia is most prevalent. Response to agronomic biofortification varied with cultivar, Mg-fertiliser rate, site and weather. The cost:benefit of these approaches and farmer acceptability, and the impact on cattle and sheep grazing on grasses biofortified with Mg requires further investigation.

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Identification of the valid reference genes for quantitative RT-PCR in annual ryegrass (Lolium multiflorum) under salt stress.

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