Proteomics in globe artichoke: protein extraction and sample complexity reduction by PEG fractionation.
Here, we report the first leaf proteome analysis for globe artichoke. Three protein extraction protocols were tested and a reproducible Mg/NP-40-based method was established. Ribulose-1,5-biphosphate carboxylase-oxygenase (RuBisCO) is a highly abundant leaf protein, and its presence masks co-localizing, less abundant proteins. To remove RuBisCO from the sample, and thereby improve spot resolution, a PEG fractionation approach was elaborated. 2-DE profiles of various PEG fractions showed that the fractionation procedure was successful in excluding most of the RuBisCO, allowing for the detection of many low-abundance proteins. Western blot analysis was able to confirm the reduction in RuBisCO content achieved by PEG fractionation. In all, 841 distinct protein spots were detected, and 40 of these, selected from the RuBisCO region of the 2-DE profile, were successfully identified by MS. A number of homologues of these proteins also co-localize with RuBisCO in Arabidopsis thaliana.
Acquadro
Alberto
A
DiVaPRA, Plant Genetics and Breeding, University of Torino, Grugliasco, Italy.
Falvo
Sara
S
Mila
Silvia
S
Giuliano Albo
Alessandra
A
Comino
Cinzia
C
Moglia
Andrea
A
Lanteri
Sergio
S
D015153:Blotting, Western; D005591:Chemical Fractionation / Q000379:methods*; D031943:Cynara scolymus / Q000737:chemistry*; D015180:Electrophoresis, Gel, Two-Dimensional; D013058:Mass Spectrometry; D018515:Plant Leaves / Q000201:enzymology; D010940:Plant Proteins / Q000032:analysis*; D011092:Polyethylene Glycols / Q000737:chemistry*; D020543:Proteome / Q000032:analysis*; D012273:Ribulose-Bisphosphate Carboxylase / Q000032:analysis / Q000737:chemistry
D010940:Plant Proteins; D020543:Proteome; D011092:Polyethylene Glycols; D012273:Ribulose-Bisphosphate Carboxylase