title: The effects of an ideal beta-turn on beta-2 microglobulin fold stability.
abstract: Beta-2 microglobulin (β2m) is the light chain of Class I major histocompatibility complex (MHC-I) complex. β2m is an intrinsically amyloidogenic protein capable of forming amyloid fibrils in vitro and in vivo. β2m displays the typical immunoglobulin-like fold with a disulphide bridge (Cys25-Cys80) cross-linking the two β-sheets. Engineering of the loop comprised between β-strands D and E has shown that mutations in this region affect protein structure, fold stability, folding kinetics and amyloid aggregation properties. Such overall effects have been related to the DE loop backbone structure, which presents a strained conformation in the wild-type (wt) protein, and a type I β-turn in the W60G mutant. Here, we report a biophysical and structural characterization of the K58P-W60G β2m mutant, where a Pro residue has been introduced in the type I β-turn i + 1 position. The K58P-W60G mutant shows improved chemical and temperature stability and faster folding relative to wt β2m. The crystal structure (1.25 Å resolution) shows that the Cys25-Cys80 disulphide bridge is unexpectedly severed, in agreement with electrospray ionization-mass spectrometry (ESI-MS) spectra that indicate that a fraction of the purified protein lacks the internal disulphide bond. These observations suggest a stabilizing role for Pro58, and stress a crucial role for the DE loop in determining β2m biophysical properties.
authors.affiliation: Dipartimento di Scienze Biomolecolari e Biotecnologie, CNR-INFM and CIMAINA, Università di Milano, Via Celoria 26, 20133 Milan, Italy.
authors.forename: Matteo
authors.initials: M
authors.lastname: Colombo
authors.forename: Stefano
authors.initials: S
authors.lastname: Ricagno
authors.forename: Alberto
authors.initials: A
authors.lastname: Barbiroli
authors.forename: Carlo
authors.initials: C
authors.lastname: Santambrogio
authors.forename: Sofia
authors.initials: S
authors.lastname: Giorgetti
authors.forename: Sara
authors.initials: S
authors.lastname: Raimondi
authors.forename: Francesco
authors.initials: F
authors.lastname: Bonomi
authors.forename: Rita
authors.initials: R
authors.lastname: Grandori
authors.forename: Vittorio
authors.initials: V
authors.lastname: Bellotti
authors.forename: Martino
authors.initials: M
authors.lastname: Bolognesi
mesh_terms: D000544:Alzheimer Disease / Q000378:metabolism; D019943:Amino Acid Substitution; D000682:Amyloid / Q000378:metabolism; D000686:Amyloidosis / Q000378:metabolism; D018360:Crystallography, X-Ray; D006801:Humans; D009154:Mutation; D011392:Proline / Q000737:chemistry / Q000235:genetics; D017510:Protein Folding; D055550:Protein Stability; D017433:Protein Structure, Secondary; D001613:beta 2-Microglobulin / Q000737:chemistry* / Q000235:genetics*
chemical_list: D000682:Amyloid; D001613:beta 2-Microglobulin; D011392:Proline