title: Nuclear magnetic resonance characterization of the refolding intermediate of beta2-microglobulin trapped by non-native prolyl peptide bond.
abstract: beta(2)-Microglobulin (beta2-m), a light chain of the major histocompatibility complex type I, is also found as a major component of amyloid fibrils formed in dialysis-related amyloidosis. Denaturation of beta2-m is considered to initiate the formation of fibrils. To clarify the mechanism of fibril formation, it is important to characterize the intermediate conformational states at the atomic level. Here, we investigated the refolding of beta2-m from the acid-unfolded state by heteronuclear magnetic resonance and circular dichroism spectroscopies. At low temperature, beta2-m refolded slowly, accumulating a rate-limiting intermediate with non-native chemical shift dispersions for several residues, but with compactness and secondary structures similar to those of the native protein. beta2-m has a cis proline residue at Pro32, located on the turn connecting the betaB and betaC strands. The slow refolding phase disappeared upon mutation of Pro32 to Val, indicating that Pro32 is responsible for the accumulation of the intermediate. The distribution of the perturbed residues in the intermediate suggests that the non-native prolyl peptide bond of Pro32 affects large areas of the molecule. A cis proline residue is common to various immunoglobulin domains involved in amyloidosis, implying that a non-native prolyl peptide bond that might occur under physiological conditions is related to the amyloidogenicity of these immunoglobulin domains.
authors.affiliation: Institute for Protein Research, Osaka University and CREST, Japan Science and Technology Agency, Yamadaoka 3-2, Suita, Osaka 565-0871, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Kameda
authors.forename: Masaru
authors.initials: M
authors.lastname: Hoshino
authors.forename: Takashi
authors.initials: T
authors.lastname: Higurashi
authors.forename: Satoshi
authors.initials: S
authors.lastname: Takahashi
authors.forename: Hironobu
authors.initials: H
authors.lastname: Naiki
authors.forename: Yuji
authors.initials: Y
authors.lastname: Goto
mesh_terms: D000143:Acids / Q000494:pharmacology; D002942:Circular Dichroism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007536:Isomerism; D007700:Kinetics; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D019906:Nuclear Magnetic Resonance, Biomolecular; D010455:Peptides / Q000737:chemistry; D011392:Proline / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D020673:Protein Renaturation; D017434:Protein Structure, Tertiary; D001613:beta 2-Microglobulin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D000143:Acids; D010455:Peptides; D001613:beta 2-Microglobulin; D011392:Proline