Probing the Urea Dependence of Residual Structure in Denatured Human α-Lactalbumin

Authors

Victoria A. Higman, University of Oxford
Heike I. Rösner, University of Copenhagen
Raffaella Ugolini, University of Oxford
Lesley H. Greene, Old Dominion UniversityFollow
Christina Redfield, University of Oxford
Lorna J. Smith, University of Oxford

Document Type

Article

Publication Date

2009

DOI

10.1007/s10858-009-9342-y

Publication Title

Journal of Biomolecular NMR

Volume

45

Issue

1

Pages

121-131

Abstract

Backbone ¹⁵N relaxation parameters and ¹⁵N–¹H^N residual dipolar couplings (RDCs) have been measured for a variant of human α-lactalbumin (α-LA) in 4, 6, 8 and 10 M urea. In the α-LA variant, the eight cysteine residues in the protein have been replaced by alanines (all-Ala α-LA). This protein is a partially folded molten globule at pH 2 and has been shown previously to unfold in a stepwise non-cooperative manner on the addition of urea. ¹⁵N R₂ values in some regions of all-Ala α-LA show significant exchange broadening which is reduced as the urea concentration is increased. Experimental RDC data are compared with RDCs predicted from a statistical coil model and with bulkiness, average area buried upon folding and hydrophobicity profiles in order to identify regions of non-random structure. Residues in the regions corresponding to the B, D and C-terminal 3₁₀ helices in native α-LA show R₂ values and RDC data consistent with some non-random structural propensities even at high urea concentrations. Indeed, for residues 101–106 the residual structure persists in 10 M urea and the RDC data suggest that this might include the formation of a turn-like structure. The data presented here allow a detailed characterization of the non-cooperative unfolding of all-Ala α-LA at higher concentrations of denaturant and complement previous studies which focused on structural features of the molten globule which is populated at lower concentrations of denaturant.

Rights

© The Authors 2009.

This is an open access article distributed under the terms of the Creative Commons Attribution Noncommercial 2.0 Generic (CC BY-NC 2.0) License, which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

Original Publication Citation

Higman, V. A., Rösner, H. I., Ugolini, R., Greene, L. H., Redfield, C., & Smith, L. J. (2009). Probing the urea dependence of residual structure in denatured human α-lactalbumin. Journal of Biomolecular NMR, 45(1), 121-131. https://doi.org/10.1007/s10858-009-9342-y

Repository Citation

Higman, Victoria A.; Rösner, Heike I.; Ugolini, Raffaella; Greene, Lesley H.; Redfield, Christina; and Smith, Lorna J., "Probing the Urea Dependence of Residual Structure in Denatured Human α-Lactalbumin" (2009). Chemistry & Biochemistry Faculty Publications. 335.
https://digitalcommons.odu.edu/chemistry_fac_pubs/335