title: Protein interactomics based on direct molecular fishing on paramagnetic particles: experimental simulation and SPR validation.
abstract: We describe an experimental approach for direct molecular fishing of prey protein on the surface of two types of paramagnetic particles (PMP) having different size and composition. Human microsomal cytochrome b(5) (b(5)) and its known partner human cytochrome P450 3A5 (CYP3A5) were used as bait and prey proteins, respectively. For assessing the level of unspecific binding of background proteins, α-fetoprotein (aFP) was used. SPR measurements were applied for quantitative analysis of trapped proteins (CYP3A5 and aFP) after fishing on PMP. It was shown that the described approach of molecular fishing on micro-PMP provides enough prey proteins for LC-MS/MS identification and SPR validation, so this approach can be used for discovery of new protein-protein interactions in the framework of Human Proteome Project.
authors.affiliation: Orechovich Institute of Biomedical Chemistry of RAMS, Moscow, Russia.
authors.forename: Pavel
authors.initials: P
authors.lastname: Ershov
authors.forename: Yury
authors.initials: Y
authors.lastname: Mezentsev
authors.forename: Oksana
authors.initials: O
authors.lastname: Gnedenko
authors.forename: Dmitry
authors.initials: D
authors.lastname: Mukha
authors.forename: Aleksei
authors.initials: A
authors.lastname: Yantsevich
authors.forename: Vladimir
authors.initials: V
authors.lastname: Britikov
authors.forename: Leonid
authors.initials: L
authors.lastname: Kaluzhskiy
authors.forename: Evgeny
authors.initials: E
authors.lastname: Yablokov
authors.forename: Andrey
authors.initials: A
authors.lastname: Molnar
authors.forename: Alexis
authors.initials: A
authors.lastname: Ivanov
authors.forename: Andrey
authors.initials: A
authors.lastname: Lisitsa
authors.forename: Andrey
authors.initials: A
authors.lastname: Gilep
authors.forename: Sergey
authors.initials: S
authors.lastname: Usanov
authors.forename: Alexander
authors.initials: A
authors.lastname: Archakov
mesh_terms: D051544:Cytochrome P-450 CYP3A / Q000737:chemistry / Q000378:metabolism; D015786:Cytochromes b5 / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D058185:Magnetite Nanoparticles / Q000737:chemistry; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D020349:Surface Plasmon Resonance / Q000379:methods* / Q000592:standards
chemical_list: D058185:Magnetite Nanoparticles; D011506:Proteins; D015786:Cytochromes b5; C510164:CYP3A5 protein, human; D051544:Cytochrome P-450 CYP3A