title: Recovering corn germ enriched in recombinant protein by wet-fractionation.
abstract: Corn wet-fractionation processes (quick-germ fractionation and traditional wet milling) were evaluated as means of recovering fractions rich in recombinant collagen-related proteins that were targeted for expression in the germ (embryo) of transgenic corn. Transgenic corn lines accumulating a recombinant full-length human collagen type-I-alpha-1 (full-length rCIalpha1) or a 44-kDa rCIalpha1 fragment targeted for seed expression with an embryo-specific promoter were used. Factors to consider in efficient recovery processes are the distribution of the peptides among botanical parts and process recovery efficiency. Both recombinant proteins were distributed 62-64% in germ comprising about 8.6% of the dry grain mass; 34-38% in the endosperm comprising 84% of the dry grain mass; 1.7% in the pericarp comprising about 5% of the dry mass; and 1% in the tip-cap comprising 1.5-2% of the dry mass. The quick-germ method employed a short steeping period either in water or SO(2)-lactic acid solution followed by wet-milling degermination to recover a germ-rich fraction. Of the total recombinant protein expressed in germ, the quick-germ process recovered 40-43% of the total recombinant protein within 6-8% of the corn mass. The traditional corn wet-milling process produced higher purity germ but with lower recovery (24-26%) of the recombinant protein. The two quick-germ methods, using water alone or SO(2)-lactic acid steeping, did not substantially differ in rCIalpha1 recovery, and the quick-germ processes recovered germ with less leaching and proteolytic losses of the recombinant proteins than did traditional wet milling. Thus, grain fractionation enriched the recombinant proteins 6-fold higher than that of unfractionated kernels. Such enrichment may improve downstream processing efficiency and enable utilizing the protein-lean co-products to produce biofuels and biorenewable chemicals by fermenting the remaining starch-rich fractions.
authors.affiliation: Center for Crops Utilization Research, Department of Food Science and Human Nutrition, Iowa State University, Ames, IA 50011-1061, USA.
authors.forename: Ilankovan
authors.initials: I
authors.lastname: Paraman
authors.forename: Steven R
authors.initials: SR
authors.lastname: Fox
authors.forename: Matthew T
authors.initials: MT
authors.lastname: Aspelund
authors.forename: Charles E
authors.initials: CE
authors.lastname: Glatz
authors.forename: Lawrence A
authors.initials: LA
authors.lastname: Johnson
mesh_terms: D005591:Chemical Fractionation / Q000379:methods*; D000090602:Collagen Type I, alpha 1 Chain; D030821:Plants, Genetically Modified / Q000737:chemistry*; D011994:Recombinant Proteins / Q000302:isolation & purification*; D012639:Seeds / Q000737:chemistry*; D017155:Wettability; D003313:Zea mays / Q000235:genetics* / Q000378:metabolism*
chemical_list: C000716127:COL1A1 protein, human; D000090602:Collagen Type I, alpha 1 Chain; D011994:Recombinant Proteins