Disulphide structure of a sunflower seed albumin: conserved and variant disulphide bonds in the cereal prolamin superfamily. FEBS Letters 396, 285-288
ABSTRACT Disulphide mapping of a methionine-rich 2S albumin from sunflower seeds showed four intra-chain disulphide bonds which are homologous with those in a related heterodimeric albumin from lupin seeds (conglutin delta). Similar conserved disulphide bonds are also present in alpha-gliadin and gamma-gliadin storage proteins of wheat, but a lower level of conservation is present in a further related group of proteins, the cereal inhibitors of alpha-amylase and trypsin. These differences may relate to the different functions of the proteins.
Full-textDOI: · Available from: Arthur Tatham, Aug 31, 2015
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- "Formed by the oxidation of cysteine-thiol groups, they contribute to the stabilization of the threedimensional structure of proteins. Cysteine bridges of sunflower albumins have been mapped (Egorov et al., 1996) but those of the globulins have not yet been directly studied. As most of the dicotyledonous globulins have similar structures (Marcone et al., 1998), their concentration must be close to those measured for linseed globulins, that is to say 61.4 µmol/g of proteins (Li-Chan and Ma, 2002). "
ABSTRACT: Through a twin-screw extrusion process the native structure of sunflower oil cake was completely transformed (globular protein denaturation/texturization and husk fiber defibration) into a simpler matrix-fiber structure, as could be seen on SEM micrographs. Further chemical reduction of protein disulfide bridges greatly reduced the melt viscosity of the moistened composite that it could be injection-molded. The molded specimens were tested and their tensile and flexural properties and water absorption calculated. Their water resistance appeared to be particularly high, and could be enhanced further after a thermal treatment (N2, 200 degrees C). The proteic matrix seemed to behave like a natural thermoset resin. Sunflower oil cake could be used without any additives to make biodegradable, water resistant and exceptionally cheap materials.Bioresource Technology 04/2006; 97(4):553-61. DOI:10.1016/j.biortech.2005.04.022 · 5.04 Impact Factor
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- "PSC33 has two more cysteines than the other 2S albumins. The Wrst one, C84, is also found at the same position in lupin conglutin and WAI 0.19, 0.28, and 0.53 . In conglutin it is unpaired whereas in WAIs it is paired with C28 and C29. "
ABSTRACT: In this work, we describe the expression, purification, and disulfide mapping of the named 'peanut seed cDNA 33' (PSC33) peanut allergen. A variant of PSC33 (with N(63), E(64), Q(69) instead of D(63), Q(64), E(69)) has been identified in peanut by proteomic analysis of a highly IgE immunoreactive purification fraction. It is 92% homologous to Ara h 6. We raised monoclonal antibodies against PSC33 and amplified it by PCR from peanut leaf genomic DNA. PSC33 was intron-less and the two NEQ and DQE variants of PSC33 were equally amplified. Since expression of the natural PSC33 (DQE) gene was very low in Escherichia coli even with supplementation of rare codon tRNAs, a synthetic gene optimized for expression in E. coli of PSC33 (DQE) was introduced into a pET9-c vector. A high production of protein occurred in the inclusion bodies that was submitted to refolding using an additive-introduced stepwise dialysis protocol which consists in the gradual removal of the denaturing agent guanidine-HCl with controlled introduction of oxidized and reduced glutathione and l-arginine as a chemical chaperone. After reverse phase HPLC purification, 1mg of pure refolded protein (as assayed by MALDI-TOF mass spectrometry, mouse IgG immunoreactivity and circular dichroism) were obtained with every 100ml of bacterial culture. Trypsin and CNBr hydrolysis of the protein combined with MALDI-TOF mass spectrometry allowed us to assign disulfide bridges and show that the native and refolded proteins were identical. The four disulfides of canonical 2S albumins were conserved and the two supplementary cysteines of PSC33 were paired together.Protein Expression and Purification 01/2006; 44(2):110-20. DOI:10.1016/j.pep.2005.05.015 · 1.51 Impact Factor
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- "SFA8 was the quantitatively major component of the two and has subsequently been studied in some detail. This includes determination of its amino acid sequence (Kortt et al., 1991), disulphide structure (Egorov et al., 1996) folding and structure (Pandya et al., 1999, 2000). A cDNA clone has also been isolated (Kortt et al., 1991) and used to increase the methionine content of transgenic lupin seeds (Molvig et al., 1997). "
ABSTRACT: 2S albumin fractions were isolated by a modified acetone precipitationmethod (Kortt and Caldwell 1990) from seeds of 103 sunflower (Helianthus annuus L.) accessions and analysed by SDS-PAGE, IEF andRP-HPLC. Two methionine-rich albumins SFA7 and SFA8 showed nodifferences in mobility on SDS-PAGE gels but were readily separated byRP-HPLC. Their levels also varied widely between different genotypes, inrelation to each other and as proportions of the total albumin fraction. Avariant form of SFA8 was identified which differed from the normal SFA8in its pI (6.5 compared to 6.0) and mobility on SDS-PAGE. N-terminal sequences of both the variant form of SFA8 and the majorform of SFA7 were identical to that reported previously for the normalform of SFA8 from the cultivar Hysun (Kortt et al., 1991) indicating theirstructural relatedness. Analysis of segregation in the F2 of the crossbetween lines VIR130 (variant SFA8) and VIR104 (normal SFA8) showedthat the normal and variant forms of SFA8 are encoded by alleles at asingle Mendelian locus. The levels of SFA7 and SFA8 in the seeds ofparental lines, F1 hybrids and individual F2 seeds classifiedfrom SDS-PAGE and IEF as homozygous for normal SFA8 (VIR104 type),homozygous for variant SFA8 (VIR130 type) and heterozygous (F1type) were determined by RP-HPLC. Seeds of the parental line VIR130contained 3.7% SFA7 and 19.0% SFA8 whereas seeds of VIR104contained 9.9% SFA7 and 12.8% SFA8. The F1 hybrid seedscontained a higher total amount of SFA7+8 proteins (32% comparingto 22% in each parent) which was largely accounted for by a highproportion of SFA7. The mean combined proportions of SFA7+8 in eachof the three phenotypic classes of F2 seeds were about 18–19% ofthe total. However, the combined proportions of SFA7+8 varied in therange 10–20% among the individual seeds. The ratio of SFA7 to SFA8was highest in the VIR104-type and heterozygous seeds, with the amountof SFA7 exceeding that of SFA8 in six heterozygous seeds. Theproportions of SFA7 and SFA8 were inversely correlated among individualF2 seeds. The results suggest that the amounts and proportions ofSFA7 and SFA8 are determined by genetic factors in addition toavailability of sulphur.Euphytica 12/2002; 129(1):99-107. DOI:10.1023/A:1021562712945 · 1.69 Impact Factor