Structural characterization of the molten globule and native states of ovalbumin: a 1H NMR study.

Molecular characteristics of ovalbumin (OVA) in the acidic (pD 3.08, the E-form) and neutral [pD 7.29, the N-form (native form)] regions were studied by measuring effective radii, 1H NMR spectra, spin-echo 1H NMR spectra and cross-relaxation times (TIS) from irradiated to observed protein protons which are particularly sensitive for detection of the mobile segments and/or structural looseness in proteins. 1H NMR spectra did not show significant differences between the N- and E-forms except for the spectral lines in the CH3, epsilon CH2 and aromatic regions. Effective radii and TIS values for main- and side-chains showed 1.08 and 1.5- to 2.0-fold increases on going from the N- to E-forms, respectively. The elongation of TIS values might indicate the appearance of the fluctuating tertiary structure in the E-form. Molecular characteristics of the E-form, inferred from reported far ultraviolet-circular dichroism (UV-CD) spectra in the peptide region, near UV-CD spectra in the aromatic region [Koseki et al. (1988) J. Biochem. 103, 425-430], effective radii and especially elongation of TIS values might indicate that the E-form could be in the molten globule state. The onset of denaturation of OVA using TIS measurements was also studied.