Characterization and modelling of the hydrophobic domain of a sunflower oleosin.
The oleosins are a group of hydrophobic proteins present on the surface of oil bodies in seeds, where they are thought to prevent coalescence. They contain a central hydrophobic domain of 68-74 residues that is thought to form a loop into the triacylglycerol matrix of the oil body, but the conformation adopted by this sequence is uncertain. We have therefore expressed an oleosin cDNA from sunflower (Helianthus annuus L.) in Escherichia coli as a fusion with maltose-binding protein (MBP) and isolated a peptide corresponding to the hydrophobic domain by sequential digestion with factor Xa (to remove the MBP) followed by trypsin and Staphylococcus V8 protease to remove the N- and C-terminal domains of the oleosin. Circular dichroism spectroscopy of the peptide in two solvent systems chosen to mimic the environment within the oil body (trifluoroethanol and SDS) demonstrated high proportions of alpha-helical structure, with no beta-sheet. A model was therefore developed in which the domain forms an alpha-helical hairpin structure, the two helices being separated by a turn region. We consider that this model is consistent with our current knowledge of oleosin structure and properties.
Alexander
Lucille G
LG
IACR-Long Ashton Research Station, Department of Agricultural Sciences, University of Bristol, UK.
Sessions
Richard B
RB
Clarke
Anthony R
AR
Tatham
Arthur S
AS
Shewry
Peter R
PR
Napier
Johnathan A
JA
D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002942:Circular Dichroism; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D006368:Helianthus / Q000235:genetics* / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D058619:Maltose-Binding Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D002352:Carrier Proteins; D018076:DNA, Complementary; D058619:Maltose-Binding Proteins; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; C079350:oleosin, Helianthus annuus