Purification and activity of a wheat 9-kDa lipid transfer protein expressed in Escherichia coli as a fusion with the maltose binding protein.
The cDNA encoding a wheat (Triticum durum) lipid transfer protein of 9 kDa was inserted into an Escherichia coli expression vector, pIH902, and expressed in the bacteria as a fusion with the maltose binding protein. The fusion protein was then purified to homogeneity and subjected to factor Xa cleavage. Although complete cleavage of the fusion protein was obtained, the expected lipid transfer protein was not recovered; it appears to be degraded during protease digestion. However, a fluorescent lipid transfer assay demonstrated that the fusion protein has an activity identical to that of the wheat-purified lipid transfer protein. Thus, this expression system should allow further understanding of the structure/function relationships of wheat lipid transfer proteins.
Dieryck
W
W
Unité de Biochimie et Biologie Moléculaire des Céréales, INRA, Montpellier, France.
Lullien-Pellerin
V
V
Marion
D
D
Joudrier
P
P
Gautier
M F
MF
D018528:ATP-Binding Cassette Transporters; D052179:Antigens, Plant; D001483:Base Sequence; D001692:Biological Transport; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification*; D003001:Cloning, Molecular / Q000379:methods; D018076:DNA, Complementary; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D015951:Factor Xa / Q000737:chemistry / Q000378:metabolism; D005822:Genetic Vectors / Q000737:chemistry / Q000235:genetics; D050356:Lipid Metabolism; D058619:Maltose-Binding Proteins; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification*; D014908:Triticum / Q000737:chemistry*
D018528:ATP-Binding Cassette Transporters; D052179:Antigens, Plant; D002352:Carrier Proteins; D018076:DNA, Complementary; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; C511920:lipid transfer proteins, plant; C461599:maltose transport system, E coli; D015951:Factor Xa