Human (Homo sapiens)
Beta2-microglobulin (Beta2-m) is a light chain of the major histocompatibility complex class I (MHC-I). Beta2-m is associated with the heavy chain through multiple hydrogen bonds, and also important for cell surface expression of MHC I.
Beta2-m is continuously shed from the cell membrane and then completely catabolized in the kidney. However, in patients undertaking long-term haemodialysis, the concentration of Beta2-m in plasma increases up to 60-fold (30–50 mg/ml). Under such pathological conditions, beta2-m assembles into amyloid fibrils in the joints and bones, causing a variety of arthropathies and pathological fractures, known as dialysis-related amyloidosis (DRA). The elevated concentrations of beta2-m are thought to be one of the factors for the onset of DRA. Similarly, increased concentrations of beta2-m in serum and urine have been found in patients with various diseases such as renal diseases, malignancies, infectious diseases, and Wilson's disease.
Although beta2-m has Trp residues at positions 60 and 95, both are located near the surface of the domain. Therefore, beta2-m does not have a conserved Trp common to other immunoglobulin domains, which is buried in close proximity to the disulfide bond.
To study the structure of amyloid fibrils in relation to their native fold, the authors prepared the Trp mutant and determined its structure. Trp60 and Trp95 were both replaced with Phe, and a single Trp was introduced at 39, the position corresponding to the conserved Trp, i.e., the triple mutations (W60F/W95F/L39W). In this study, the author showed that mutagenesis of Trp would be a promising tool with which to probe the mechanism by which amyloid fibrils form without disturbing the native conformations.
From above line: wild type at pH7.0, wild type at pH5.6, and mutant at pH5.6
Beta2-m comprised of 99 amino acid residues has a seven-stranded beta-sandiwich fold which is a typical immunoglobulin superfamily domain fold. Residues involved in the beta-strands include beta-strand A (7–12), beta-strand B (22–30), beta-strand C (36–41), beta-strand C' (43–44), beta-strand D (51–56), beta-strand E (62–69), beta-strand F (78–83) and beta-strand G (91–94). The two beta-sheets are stabilized by a intramolecular disulfide bond, highly conserved, between Cys25 (beta-strand B) and Cys80 (beta-strand F) and hydrophobic residues between the layers.
To compare the structures of the beta-strand D at the different pH values and conformational states, the authors first determined the structure of the isolated beta2-m monomer at pH 7.0, and then compared that to the isolated and the heavy chain-bound beta2-m structures at pH 5.7. The overall structures of the three beta2-m molecules were similar one another. The most striking difference is the disappearance of the beta-bulge in beta-strand D observed in the complex states at pH 5.7. The isolated beta2-m has one long continuous beta-strand D encompassing residues 51–56 at both pH 7.0 values. This suggests that the conversion of the beta-bulge in strand D into a contiguous beta-strand is not unique to the crystals formed under slightly acidic conditions, i.e., pH 5.7. The authors conclude that the enhanced fibrillation is more directly coupled with the decreased stability leading to the increased propensity of exposing amyloidogenic regions.
In addition, the authors also proposed deprotonation of Tyr 67 with a highly decreased pKa value, and that the increased pKa of His51 might be coupled with the decreased pKa value of Tyr67. On the other hand, the calculated B-factor values which can reflect the dynamic structures increased in the loop regions (AB, CD, EF and GH loops) and the N- and C-terminal regions. However, BC, FG and DE loops did not exhibit a notable increase in the B-factors.
Despite the triple mutations (W60F/W95F/L39W), the crystal structure of mutant beta2-m was very similar to that of wild-type beta2-m. The RMS deviations between them are 0.51 angstrom using main chain atoms, N, C-alpha, C, and O. Slight differences exist at residues Arg45-His51 on the CD loop and Lys58-Trp60 on the DE loop. The difference in the CD loop is caused by a difference in crystal packing interactions with symmetry-related beta2-m molecules. For the DE loop, the mutation of Trp to Phe should yield a void, possibly producing the deviation. The structure of mutant beta2-m shows tight contact between Trp39 and the disulfide bond, with the aromatic ring plane facing the disulfide bond in parallel as observed for other immunoglobulin domains. The results reveal the native fold of mutant beta2-m and furthermore the proximity of Trp39 to the disulfide bond, proposing the structural basis for the complete quenching of Trp fluorescence.
Protein Data Bank (PDB)
author: Young-Ho Lee