Large scale purification of rapeseed proteins (Brassica napus L.).
Rapeseed (Brassica napus L.) cruciferin (12S globulin), napin (2S albumin) and lipid transfer proteins (LTP) were purified at a multi-g scale. The procedure developed was simple, rather fast and resolutive; it permitted the recovery of these proteins with a good yield, such as 40% for cruciferin and 18% for napin. Nanofiltration eliminated the major phenolic compounds. The remaining protein fraction was fractionated by cation exchange chromatography (CEC) on a streamline SP-XL column in alkaline conditions. The unbound neutral cruciferin was polished by size exclusion chromatography. The alkaline napin isoforms and LTP, adsorbed on the beads, were eluted as a whole fraction and further separated by an other CEC step at acidic pH. Napins were polished by hydrophobic interaction chromatography (HIC). The fractions were characterized by reverse phase HPLC, electrophoresis, N-terminal sequencing and mass spectrometry. All the fractions contained less than 5% of impurities.
Bérot
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S
Unité de Recherche sur les Protéines Végétales et leurs Interactions, INRA, BP 71627, Rue de la Géraudière, F- 44316 Nantes Cedex 3, France. berot@nantes.inra.fr
Compoint
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JP
Larré
C
C
Malabat
C
C
Guéguen
J
J
D055516:2S Albumins, Plant; D000485:Allergens; D000595:Amino Acid Sequence; D052179:Antigens, Plant; D029688:Brassica napus / Q000737:chemistry*; D002352:Carrier Proteins / Q000302:isolation & purification*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D010940:Plant Proteins / Q000302:isolation & purification*; D020033:Protein Isoforms / Q000302:isolation & purification; D055314:Seed Storage Proteins; D014461:Ultracentrifugation
D055516:2S Albumins, Plant; D000485:Allergens; D052179:Antigens, Plant; D002352:Carrier Proteins; D010940:Plant Proteins; D020033:Protein Isoforms; D055314:Seed Storage Proteins; C059777:cruciferin protein, Brassica napus; C511920:lipid transfer proteins, plant; C050363:napin protein, Brassica napus