Characterization of Cu2+-binding modes in the prion protein by visible circular dichroism and multivariate curve resolution
J. B. Pollock, P.J. Cutler, J.M. Kenney, P.J. Gemperline, C.S. Burns
Visible circular dichroism (CD) spectra from the copper(II) titration of the metal-binding region of the prion protein, residues 57–98, were analyzed using the self-modeling curve resolution method multivariate curve resolution–alternating least squares (MCR-ALS). MCR-ALS is a set of mathematical tools for estimating pure component spectra and composition profiles from mixture spectra. Model-free solutions (e.g., soft models) are produced under the assumption that pure component profiles should be nonnegative and unimodal. Optionally, equality constraints can be used when the concentration or spectrum of one or more species is known. MCR-ALS is well suited to complex biochemical systems such as the prion protein which binds multiple copper ions and thus gives rise to titration data consisting of several pure component spectra with overlapped or superimposed absorption bands. Our study reveals the number of binding modes used in the uptake of Cu2+ by the full metal-binding region of the prion protein and their relative concentration profiles throughout the titration. The presence of a non-CD active binding mode can also be inferred. We show that MCR-ALS analysis can be initialized using empirically generated or mathematically generated pure component spectra. The use of small model peptides allows us to correlate specific Cu2+-binding structures to the pure component spectra.
Circular dichroism, Coordination chemistry, Ligand binding, Biochemistry