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A New Approach to the Study of Protein−Protein Interaction by FTIR: Complex Formation between Cytochrome P450BM-3 Heme Domain and FMN Reductase Domain
We describe a new approach to the study of protein−protein interaction using Fourier transform infrared spectroscopy (FTIR). This approach is based on the combination of FTIR technique with both protein titration experiments and the principal component analysis (factor analysis) of the IR absorption...
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Published in: | Biochemistry (Easton) 2002-11, Vol.41 (46), p.13514-13525 |
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Main Authors: | , , , |
Format: | Article |
Language: | English |
Subjects: | |
Citations: | Items that this one cites Items that cite this one |
Online Access: | Get full text |
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Summary: | We describe a new approach to the study of protein−protein interaction using Fourier transform infrared spectroscopy (FTIR). This approach is based on the combination of FTIR technique with both protein titration experiments and the principal component analysis (factor analysis) of the IR absorption spectra in the 1500−1800 cm-1 region for the protein mixtures. We have applied this approach to the interaction of the heme domain with the FMN domain of bacterial monooxygenase cytochrome P450BM-3 (CYP102A1). The analysis reveals that the first principal component reflects the protein−protein complex formation because the loading factors show a clear systematic dependence on the concentration of the heme domain according to a titration curve with a dissociation constant of ∼5 μM. The spectrum of the first principal component has been assigned to structural changes in the secondary structure (increase of β-sheet and α-helix and decrease of turn structures), amino acid side chains (protonation of aspartate and C-terminal COO group), and deprotonation of a propionic acid COOD group in the heme. |
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ISSN: | 0006-2960 1520-4995 |
DOI: | 10.1021/bi0262505 |