NMR characterization of the interaction between cytochrome c552 and the CuA-soluble fragment of the ba3 oxidase from Thermus thermphilus

ALEJANDRO J. VILA; JAVIER MARCELO GONZALEZ; CLAUDIO FERNANDEZ

Congreso; XXXIX Reunión Anual de la Sociedad Argentina de Biofísica; 2003

The structural analysis of protein-protein interactions is one of the most challenging tasks nowadays in structural biology. The protein-protein (transient) complexes between the various components of the respiratory chain are characterized by a lifetime on the order of milliseconds and KD´s in the milimolar to micromolar range, thus complicating the  structural analysis of the complexes. Chemical shift mapping by NMR spectroscopy is a useful tool to investigate such transient contacts, since it can monitor changes in the electron-shielding properties of a protein as the result of temporary contacts with a reaction partner. In this study, we investigated the molecular interaction between two components of the electron-transfer chain from Thermus thermophilus: the engineered, water-soluble fragment of cytochrome c552 and the CuA domain from cytochrome oxidase. Comparison of 1H-15N HSQC spectra of the [15N]-labeled CuA fragment in the absence and presence of the cytochrome c552 fragment showed chemical shift changes for the backbone amide groups of a discrete number of resonances. The mapped contact areas on the CuA fragment surface were comparable in the presence of both reduced and oxidized cytochrome c552 species, suggesting that the respective chemical shits changes represent biologically relevant  protein-protein interactions.