A versatile method for producing labeled or unlabeled Aβ55, Aβ40, and other β-amyloid family peptides

Nanodroplet Oligomers (NanDOs) of Aβ40

“…We expressed the Aβ40 peptide, both unlabeled and labeled, for NMR spectroscopy as reported elsewhere . Briefly, Aβ40 was expressed as an N-terminal hexa-His tag appended to Aβ40 by a Factor Xa cleavage site using a pET 28a vector.…”

“…The peptide was dissolved in a small volume of a 50/50/0.1 (v/v/v) water/acetonitrile/TFA mixture and lyophilized. Peptides were purified by RP-HPLC as described previously . When the isoacyl dipeptide strategy had been used, following RP-HPLC purification and lyophilization, the iso–acyl bond was converted to a peptide bond by dissolving it in 0.1 M NH 4 HCO 3 (to obtain a pH of ≥7) and incubated for 10 min, after which the Aβ peptides were repurified by RP-HPLC.…”

“…We expressed the Aβ40 peptide, both unlabeled and labeled, for NMR spectroscopy as reported elsewhere. 32 Briefly, Aβ40 was expressed as an N-terminal hexa-His tag appended to Aβ40 by a Factor Xa cleavage site using a pET 28a vector. Aβ40 is well expressed in minimal medium containing either 14 N or 15 N in NH 4 Cl as the sole nitrogen source.…”

“…Peptide synthesis of Aβ40 (NH 2 -DAEFRHDSGYEVHHQ-KLVFFAEDVGSNKGAIIGLMVGGVV-COOH), and of the same peptide with either Y10 or L34 mutated to C, was carried out using an Applied Biosystems (Foster City, CA) 433A synthesizer, essentially as described in an earlier publication. 32 In general, Aβ40 was synthesized using the dipeptide Boc-Ser(Fmoc-Gly)-OH (Novabiochem) for G25-S26. 33 Standard Fmoc synthesis was carried out from residue V40 to N27.…”