A pH-dependent conformational transition of Aβ peptide and physicochemical properties of the conformers in the glial cell

Abstract

In the present study we identified the epitopes of antibodies against amyloid β-(1–42)-peptide (Aβ1–42): 4G8 reacted with peptides corresponding to residues 17–21, 6F/3D reacted with peptides corresponding to residues 9–14, and anti 5-10 reacted with peptides corresponding to residues 5–10. The study also yielded some insight into the Aβ1–42 structures resulting from differences in pH. An ELISA study using monoclonal antibodies showed that pH-dependent conformational changes occur in the 6F/3D and 4G8 epitopes modified at pH 4.6, but not in the sequences recognized by anti 1-7 and anti 5-10. This was unique to Aβ1–40 and Aβ1–42 and did not occur with Aβ1–16 or Aβ17–42. The reactivity profile of 4G8 was not affected by blockage of histidine residues of pH-modified Aβ1–40 and Aβ1–42 with diethyl pyrocarbonate; however, the mutant [Gln11]Aβ1–40 abrogated the unique pH-dependence towards 4G8 observed with Aβ1–40. These findings suggest that these epitopes are cryptic at pH4.6, and that Glu11 is responsible for the changes. We suggest that the abnormal folding of 6F/3D epitope affected by pH masked the 4G8 epitope. A study of the binding of metal ions to Aβ1–42 suggested that Cu2+ and Zn2+ induced a conformational transition around the 6F/3D region at pH7.4, but did not affect the region when it was modified at pH4.6. However, Fe2+ had no effect, irrespective of pH. Aβ modified at pH 4.6 appeared to be relatively resistant to proteinase K compared with Aβs modified at pH7.4, and the former might be preferentially internalized and accumulated in a human glial cell. Our findings suggest the importance of microenvironmental changes, such as pH, in the early stage of formation of Aβ aggregates in the glial cell.

Immunohistochemical Visualization of Amyloid-β Protein Precursor and Amyloid-β in Extra- and Intracellular Compartments in the Human Brain

“…Clone 6F3D is raised against Aβ 8−17 (DakoCytomation) [3,58]. Clone 4G8 (Signet) is raised against Aβ17-24 (Signet) [3,58]. Clone 12F4 is raised against Aβ 1−42 (Covance) and is reactive to C-terminus of Aβ and is specific for the isoform ending at the 42nd amino acid.…”

“…Clone 6E10 is raised against Aβ 1−17 (Signet) [3,56,57]. Clone 6F3D is raised against Aβ 8−17 (DakoCytomation) [3,58]. Clone 4G8 (Signet) is raised against Aβ17-24 (Signet) [3,58].…”

Different Immunoreactivity against Monoclonal Antibodies between Wild-type and Mutant Copper/Zinc Superoxide Dismutase Linked to Amyotrophic Lateral Sclerosis

“…A conformational transition of the prion protein has been detected by various mAbs (24 -26). The pH-dependent conformational transition of the Alzheimer ␤-amyloid peptide was monitored by their mAbs (27). Inhibition of the fibrillar aggregation of the Alzheimer ␤-amyloid peptide (28) and inhibition of prion transmission (29) have also been reported, based on the use of mAbs.…”

Discrete conformational changes as regulators of the hydrolytic properties of beta‐amyloid (1–40)

“…Reverse structure Abeta , shorter fragments of the peptide (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35) and two unrelated peptides (bradykinin and bombesin) did not cause hydrolysis of fluorescein dibutyrate (supplementary Fig. S1).…”

“…An ELISA study using mAbs showed that pH-dependent conformational changes occur in the region 10-24 [30]. This region comprises charged amino acids of pH-dependent protonation that may be involved in the peptide conformational changes.…”