Mimmi Maria C, Jørgensen Thomas J D, Pettirossi Fabio, Corazza Alessandra, Viglino Paolo, Esposito Gennaro, De Lorenzi Ersilia, Giorgetti Sofia, Pries Mette, Corlin Dorthe B, Nissen Mogens H, Heegaard Niels H H
Dipartimento di Scienze e Tecnologie Biomediche and MATI Centre of Excellence, University of Udine, Italy.
FEBS J. 2006 Jun;273(11):2461-74. doi: 10.1111/j.1742-4658.2006.05254.x.
Cleavage of the small amyloidogenic protein beta2-microglobulin after lysine-58 renders it more prone to unfolding and aggregation. This is important for dialysis-related beta2-microglobulin amyloidosis, since elevated levels of cleaved beta2-microglobulin may be found in the circulation of dialysis patients. However, the solution structures of these cleaved beta2-microglobulin variants have not yet been assessed using single-residue techniques. We here use such methods to examine beta2-microglobulin cleaved after lysine-58 and the further processed variant (found in vivo) from which lysine-58 is removed. We find that the solution stability of both variants, especially of beta2-microglobulin from which lysine-58 is removed, is much reduced compared to wild-type beta2-microglobulin and is strongly dependent on temperature and protein concentration. 1H-NMR spectroscopy and amide hydrogen (1H/2H) exchange monitored by MS show that the overall three-dimensional structure of the variants is similar to that of wild-type beta2-microglobulin at subphysiological temperatures. However, deviations do occur, especially in the arrangement of the B, D and E beta-strands close to the D-E loop cleavage site at lysine-58, and the experiments suggest conformational heterogeneity of the two variants. Two-dimensional NMR spectroscopy indicates that this heterogeneity involves an equilibrium between the native-like fold and at least one conformational intermediate resembling intermediates found in other structurally altered beta2-microglobulin molecules. This is the first single-residue resolution study of a specific beta2-microglobulin variant that has been found circulating in dialysis patients. The instability and conformational heterogeneity of this variant suggest its involvement in beta2-microglobulin amyloidogenicity in vivo.
小淀粉样蛋白β2-微球蛋白在赖氨酸-58处裂解后更易于展开和聚集。这对于与透析相关的β2-微球蛋白淀粉样变性很重要，因为在透析患者的循环系统中可能会发现裂解的β2-微球蛋白水平升高。然而，尚未使用单残基技术评估这些裂解的β2-微球蛋白变体的溶液结构。我们在此使用此类方法来研究赖氨酸-58处裂解的β2-微球蛋白以及进一步加工的变体（在体内发现），后者去除了赖氨酸-58。我们发现，与野生型β2-微球蛋白相比，这两种变体的溶液稳定性，尤其是去除赖氨酸-58的β2-微球蛋白的稳定性大大降低，并且强烈依赖于温度和蛋白质浓度。1H-NMR光谱和通过质谱监测的酰胺氢（1H/2H）交换表明，在亚生理温度下，变体的整体三维结构与野生型β2-微球蛋白相似。然而，确实存在偏差，特别是在靠近赖氨酸-58处D-E环裂解位点的B、D和Eβ链的排列中，并且实验表明这两种变体存在构象异质性。二维NMR光谱表明，这种异质性涉及天然样折叠与至少一种构象中间体之间的平衡，该中间体类似于在其他结构改变的β2-微球蛋白分子中发现的中间体。这是对在透析患者体内循环的特定β2-微球蛋白变体进行的首次单残基分辨率研究。这种变体的不稳定性和构象异质性表明其在体内参与了β2-微球蛋白的淀粉样变性。
