fulltext.study @t Gmail

Amyloidogenecity and pitrilysin sensitivity of a lysine-free derivative of amyloid β-peptide cleaved from a recombinant fusion protein

Paper ID Volume ID Publish Year Pages File Format Full-Text
25800 43602 2006 12 PDF Available
Title
Amyloidogenecity and pitrilysin sensitivity of a lysine-free derivative of amyloid β-peptide cleaved from a recombinant fusion protein
Abstract

The progressive cerebral deposition of a 40–42 residues amyloid β-peptide (Aβ) is regarded as a major factor in the onset of the Alzheimer's disease. It has recently been shown that Aβ1–40 is cleaved by Escherichia coli pitrilysin, a homologue of insulysin, at a specific site. To facilitate the studies on a recognition mechanism of Aβ by pitrilysin, an overproduction system of Aβ1–40 as a fusion protein with E. coli RNase HI was constructed. This fusion protein was designed such that an Aβ1–40 derivative, Aβ1–40*, in which Lys16 and Lys28 of Aβ1–40 are simultaneously replaced by Ala, is attached to the C-terminus of E. coli RNase HI and Aβ1–40* is separated from RNase HI upon cleavage with lysyl endopeptidase. The fusion protein was overproduced in E. coli in inclusion bodies, solubilized and purified in the presence of guanidine hydrochloride, and cleaved by lysyl endopeptidase. Aβ1–40* was purified from the resultant peptide fragments by reverse-phase HPLC. Measurement of the far-UV CD spectra suggests that Aβ1–40* is conformationally similar to Aβ1–40. However, the thioflavin T binding assay suggests that Aβ1–40* is more amyloidogenic than Aβ1–40. Nevertheless, Aβ1–40* was cleaved by pitrilysin at the site identical to that in Aβ1–40.

Keywords
E. coli pitrilysin; Insulysin; Metallopeptidase; Amyloid β-peptide; Fusion protein; AmyloidogenecityAβ1–40, amyloid β-peptide 1–40; AD, Alzheimers's disease; CD, circular dichroism; GdnHCl, guanidine hydrochloride; HPLC, high-performance liquid chromatogr
First Page Preview
Amyloidogenecity and pitrilysin sensitivity of a lysine-free derivative of amyloid β-peptide cleaved from a recombinant fusion protein
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 122, Issue 2, 23 March 2006, Pages 186–197
Authors
, , , ,
Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering