Morphological Differences between β₂‐Microglobulin in Fibrils and Inclusion Bodies

Abstract

Over expression of proteins in E. coli frequently results in the production of inclusion bodies. Although β2‐microglobulin frequently forms fibrillar structures, our studies reveal significant differences between the protein in fibrils and inclusion bodies. This suggests that the formation of fibrils in inclusion bodies is dependent on the propensity of the protein to form fibrillar structures.

Structural Properties of Bacterial Inclusion Bodies

“…Direct structural comparison at the residue-specific resolution of IBs and amyloid fibrils of the same proteins has also been reported in the literature by NMR spectroscopy (Wasmer et al, 2009;Wang et al, 2010;Taylor et al, 2011). In particular, it has been found that IBs of the C-terminal region (residues 218 to 289) of the Podospora anserina prion ] display an identical chemical shift of HET-s(218-289) fibrils in the 13 C-13 C protondriven spin-diffusion solid-state NMR spectra.…”

“…Interestingly, IBs and fibrils were found to be characterized by similar infectivity, chemical stability, seeding competency, and by a similar H/D exchange pattern as studied by liquid-state NMR (Wasmer et al, 2009). Indeed, b2m IBs and fibrils (obtained in vitro at pH 2.5) displayed important differences in ThT binding and molecular structure and dynamics, as observed by solid-state NMR (Taylor et al, 2011). It is noteworthy that the opposite has been found in the case of β2microglobulin (b2m).…”

Methodological advances and strategies for high resolution structure determination of cellular protein aggregates

“…The analysis of HET-s (218–289) revealed that both IBs (raw and partially purified) spectra reproduce all the peaks visible for HET-s (218–289) fibrils assembled in vitro ( 107 ). In contrast, Aβ and β2-microglobulin IBs demonstrated fewer similarities, indicating more heterogeneous ensembles with more contributions from non-β-sheet conformations ( 108 , 109 ). While the increased protein content of IB samples is advantageous, the heterogeneity of these aggregates can present a challenge.…”

“…As IBs are often produced during protein overexpression in recombinant bacteria, the sample concentration required for ssNMR studies is more readily achieved. Comparative studies of IBs and fibrils using ssNMR have proven useful in characterizing the composition of IBs for proteins such as the prion-forming domain of HET-s fungal prion (HET-s 218–289), Aβ peptides, and β2-microglobulin ( 107 , 108 , 109 ). The analysis of HET-s (218–289) revealed that both IBs (raw and partially purified) spectra reproduce all the peaks visible for HET-s (218–289) fibrils assembled in vitro ( 107 ).…”