Quantitative Residue-Specific Protein Backbone Torsion Angle Dynamics from Concerted Measurement of ³<i>J</i> Couplings

Abstract

Three-bond ³<i>J</i>_C′C′ and ³<i>J</i>_HNHα couplings in peptides and proteins are functions of the intervening backbone torsion angle ϕ. In well-ordered regions, ³<i>J</i>_HNHα is tightly correlated with ³<i>J</i>_C′C′, but the presence of large ϕ angle fluctuations differentially affects the two types of couplings. Assuming the ϕ angles follow a Gaussian distribution, the width of this distribution can be extracted from ³<i>J</i>_C′C′ and ³<i>J</i>_HNHα, as demonstrated for the folded proteins ubiquitin and GB3. In intrinsically disordered proteins, slow transverse relaxation permits measurement of ³<i>J</i>_C′C′ and ³<i>J</i>_HNH couplings at very high precision, and impact of factors other than the intervening torsion angle on ³<i>J</i> will be minimal, making these couplings exceptionally valuable structural reporters. Analysis of α-synuclein yields rather homogeneous widths of 69 ± 6° for the ϕ angle distributions and ³<i>J</i>_C′C′ values that agree well with those of a recent maximum entropy analysis of chemical shifts, <i>J</i> couplings, and ¹H–¹H NOEs. Data are consistent with a modest (≤30%) population of the polyproline II region