title: Unraveling tobacco BY-2 protein complexes with BN PAGE/LC-MS/MS and clustering methods.
abstract: To understand physiological processes, insight into protein complexes is very important. Through a combination of blue native gel electrophoresis and LC-MS/MS, we were able to isolate protein complexes and identify their potential subunits from Nicotiana tabacum cv. Bright Yellow-2. For this purpose, a bioanalytical approach was used that works without a priori knowledge of the interacting proteins. Different clustering methods (e.g., k-means and hierarchical clustering) and a biclustering approach were evaluated according to their ability to group proteins by their migration profile and to correlate the proteins to a specific complex. The biclustering approach was identified as a very powerful tool for the exploration of protein complexes of whole cell lysates since it allows for the promiscuous nature of proteins. Furthermore, it searches for associations between proteins that co-occur frequently throughout the BN gel, which increases the confidence of the putative associations between co-migrating proteins. The statistical significance and biological relevance of the profile clusters were verified using functional gene ontology annotation. The proof of concept for identifying protein complexes by our BN PAGE/LC-MS/MS approach is provided through the analysis of known protein complexes. Both well characterized long-lived protein complexes as well as potential temporary sequential multi-enzyme complexes were characterized.
authors: Center for Proteomics (CFP), Groenenborgerlaan 171, B-2020 Antwerp, Belgium.
authors: Noor
authors: N
authors: Remmerie
authors: Thomas
authors: T
authors: De Vijlder
authors: Dirk
authors: D
authors: Valkenborg
authors: Kris
authors: K
authors: Laukens
authors: Koen
authors: K
authors: Smets
authors: Jilles
authors: J
authors: Vreeken
authors: Inge
authors: I
authors: Mertens
authors: Sebastien C
authors: SC
authors: Carpentier
authors: Bart
authors: B
authors: Panis
authors: Geert
authors: G
authors: De Jaeger
authors: Ronny
authors: R
authors: Blust
authors: Els
authors: E
authors: Prinsen
authors: Erwin
authors: E
authors: Witters
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods; D016000:Cluster Analysis; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D046912:Multiprotein Complexes / Q000302:isolation & purification*; D010940:Plant Proteins / Q000302:isolation & purification*; D053719:Tandem Mass Spectrometry / Q000379:methods; D014026:Nicotiana / Q000737:chemistry*
chemical_list: D046912:Multiprotein Complexes; D010940:Plant Proteins