title: [Mass-spectrometric identification of interaction sites between cytochrome P450 2B4 and NADPH cytochrome P450 reductase].
abstract: We determined the interaction sites of the cytochrome P450's protein-partners: 2B4 (d-2B4) and NADPH-cytochrome P450 of reductase (d-Fp). While in operation, these proteins are forming the complexes. We used 4-4'-dithio(bisphenyl)azide linker for non-specific covalent coupling of d-2B4 complexes with d-Fp in Emulgen-913-monomerized system. Covalently-linked peptides in this complex were identified with ESI-MS/MS. Several sites of these proteins' binding with each other were revealed. Based on them, a model of intermolecular protein interactions was created. The model includes 5 cross-linker-stabilized contact sites of d-2B4 with d-Fp involving the following peptides of d-2B4 and d-Fp: (1) d-2B4423-433 and d-Fp 102-109; (2) d-2B4324-336 and d-Fp570-585; (3) d-2B4327-336 and d-Fp452-464; (4) d-2B4 192-197 and d-Fp456-464; (5) d-2B4 134-139 and d-Fp406-425. Herein, in the latter two cases, the peptides of d-Fp are located in their inter-domain slit and stabilize protein-protein complex via nanoprobe cross-linker; therefore, the formation of d-2B4/d-Fp complexes in these sites may involve aminoacid residues d-Fp456-464 and d-Fp406-425 surrounding inter-domain slit.
authors.forename: A V
authors.initials: AV
authors.lastname: Ivanov
authors.forename: A T
authors.initials: AT
authors.lastname: Kopylov
authors.forename: V G
authors.initials: VG
authors.lastname: Zgoda
authors.forename: I Iu
authors.initials: IIu
authors.lastname: Toropygin
authors.forename: E V
authors.initials: EV
authors.lastname: Khriapova
authors.forename: Iu D
authors.initials: IuD
authors.lastname: Ivanov
mesh_terms: D001189:Aryl Hydrocarbon Hydroxylases / Q000737:chemistry*; D001665:Binding Sites; D003432:Cross-Linking Reagents / Q000737:chemistry; D000072467:Cytochrome P450 Family 2; D008958:Models, Molecular; D009251:NADPH-Ferrihemoprotein Reductase / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
chemical_list: D003432:Cross-Linking Reagents; D001189:Aryl Hydrocarbon Hydroxylases; D000072467:Cytochrome P450 Family 2; C465241:cytochrome P-450 CYP2B4 (rabbit); D009251:NADPH-Ferrihemoprotein Reductase