Two-step purification of pathogenesis-related proteins from grape juice and crystallization of thaumatin-like proteins.
Grape thaumatin-like (TL) proteins and chitinases play roles in plant-pathogen interactions and can cause protein haze in white wine unless removed prior to bottling. A two-step method is described that highly purified hundreds of milligrams of TL proteins and chitinases from two juices by strong cation exchange (SCX) and hydrophobic interaction chromatography (HIC). The method was fast and separated isoforms of TL proteins and chitinases from within the same juice, in most cases to >97% purity. The isolated proteins were identified by peptide nanoLC-MS/MS and crystallized using a high-throughput screening method. Crystals from three protein fractions produced high-resolution X-ray crystallography data.
Van Sluyter
Steven C
SC
The Australian Wine Research Institute, Glen Osmond, SA, Australia.
Marangon
Matteo
M
Stranks
Samuel D
SD
Neilson
Karlie A
KA
Hayasaka
Yoji
Y
Haynes
Paul A
PA
Menz
R Ian
RI
Waters
Elizabeth J
EJ
D002688:Chitinases / Q000737:chemistry* / Q000302:isolation & purification*; D002845:Chromatography / Q000379:methods*; D003460:Crystallization; D054884:Host-Pathogen Interactions; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D027843:Vitis / Q000737:chemistry* / Q000382:microbiology
D010940:Plant Proteins; C003427:thaumatin protein, plant; D002688:Chitinases