Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: (A) Sequences of WT Aβ 1–40 and its two variants with scrambled N-terminal 1–16 amino acids (NSCR) and with all amino acids scrambled (SCR). Protonation sites on side-chain moieties are highlighted in red. (B) Sequence of WT Aβ 1–40 with number of exchangeable hydrogens and charges. (C) Exchangeable side-chain hydrogens present in neutral forms of residues of Aβ 1–40. Protonation sites on basic side-chain marked in blue, neutral side-chain in green. Proton accepting groups are highlighted in red. Slower exchangeable side-chain hydrogens marked in dotted green.

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques: Sequencing

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: Difference in deuterium uptake per monomer between dimers and monomers (circles) and trimers and monomers (tringles) in WT Aβ 1–40 (A), NSCR Aβ 1–40 (B) and SCR Aβ 1–40 (C), based on the analysis of the m/z region 2164–2169 which contains well resolved signals from MON 2+ , DIM 4+ , TRI 6+ oligomeric forms bearing the same charge per monomer. The difference in deuterium uptake (D) is shown at the vertical axis and HDX reagent gas flow rate (20–50 mL/min) at the horizontal axis. Four batch-to-batch replicates are represented on the scatter. Dashes refer to the mean difference. Error bars indicate the standard deviation calculated from replicate measurements (number of replicates n was 4).

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques: Standard Deviation

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: Analysis of the isotopic envelopes after gas-phase HDX-MS for selected signals of species bearing the same charge per monomer, namely MON 2+ , DIM 4+ and TRI 6+ (2164–2185 m/z range) of WT Aβ 1–40 (A) and SCR Aβ 1–40 (B). Left panels—isotopic envelopes corresponding to MON 2+ (upper panels), DIM 4+ (middle panels), TRI 6+ (lower panels), for make-up gas flow 50 mL/min in the presence of ND 3 /D 2 O reagent. Right panels—decomposition of the experimental isotopic envelope into a family of isotopic envelopes of FWHM, expected for a single conformational state. The spectra were recalculated from m/z domain, (shown in left panels) to the domain of molecular mass of a monomeric unit in an oligomer (shown in right panels). Results of replicates of this experiments are shown in Figure C in .

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques:

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: Charge state distributions of MS signals of: (A) monomers, (B) dimers, (C) trimers, (D) tetramers, (E) pentamers, (F) hexamers of WT Aβ 1–40 (upper panels), NSCR Aβ 1–40 (middle panels) and SCR Aβ 1–40 (lower panels), extracted from IM-MS data, with the height of each signal equal to the maximum value of the signal in the corresponding charge envelope. In SCR, DIM 7+ was missing, DIM 6+ was less intense, and DIM 4+ was more intense. The charge state envelope was in some instances (dimers, trimers, pentamers) shifted towards a lower charge in SCR. For trimers, the dominant form was 5+ in SCR and 7+ in both WT and NSCR, while the 8+ form was missing in SCR. TET 8+ and TET 9+ were missing in SCR but the remaining 7+, 6+, and 5+ signals retained the same mutual amplitude. A major PEN 9+ signal in WT and NSCR was missing in SCR, for which PEN 7+ was predominant. Hexameric signals are absent in SCR.

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques:

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: For each selected m/z region, a profile of molecular species present at different drift times in the IM T-wave is shown, with each species assigned to a particular oligomeric charge state. Some profiles contain a single peak, indicating the presence of one form of a single collisional cross section, whereas other signals are split in the domain of the drift time into multiple species. (A) DIM 5+ —the dominant, extended form becomes more similar to the compact form in SCR. For TET 7+ (D) and TET 6+ (E), alternative structural forms of WT Aβ collapse in SCR to a single major form with intermediate drift time (and thus intermediate Ω), while TRI 7+ in SCR (B) exhibits a single form with the drift time closer to the compact form in WT. The split TRI 6+ signals as well as the TET 8+ signal (C), which are clear in WT and NSCR, become very weak in SCR. Signals were assigned to species as described before in Ref .

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques:

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: Representative autocorrelation function (ACF), directly adapted from dynamic light scattering data obtained for WT Aβ 1–40 (line) and SCR Aβ 1–40 (dashes). The autocorrelation function decays in the range of 1 down to 0, and the gap remaining to 1 observed at the shortest correlation time is indicative for the existence of monomeric or low-order oligomers, which remain undetectable. The short-correlation-time asymptote of ACF is marked by thin solid lines. The mid-point of an ACF decay is marked by the cross.

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques:

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: (A) Sequences of WT Aβ 1–40 and its two variants with scrambled N-terminal 1–16 amino acids (NSCR) and with all amino acids scrambled (SCR). Protonation sites on side-chain moieties are highlighted in red. (B) Sequence of WT Aβ 1–40 with number of exchangeable hydrogens and charges. (C) Exchangeable side-chain hydrogens present in neutral forms of residues of Aβ 1–40. Protonation sites on basic side-chain marked in blue, neutral side-chain in green. Proton accepting groups are highlighted in red. Slower exchangeable side-chain hydrogens marked in dotted green.

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques: Sequencing

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: Difference in deuterium uptake per monomer between dimers and monomers (circles) and trimers and monomers (tringles) in WT Aβ 1–40 (A), NSCR Aβ 1–40 (B) and SCR Aβ 1–40 (C), based on the analysis of the m/z region 2164–2169 which contains well resolved signals from MON 2+ , DIM 4+ , TRI 6+ oligomeric forms bearing the same charge per monomer. The difference in deuterium uptake (D) is shown at the vertical axis and HDX reagent gas flow rate (20–50 mL/min) at the horizontal axis. Four batch-to-batch replicates are represented on the scatter. Dashes refer to the mean difference. Error bars indicate the standard deviation calculated from replicate measurements (number of replicates n was 4).

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques: Standard Deviation

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: Charge state distributions of MS signals of: (A) monomers, (B) dimers, (C) trimers, (D) tetramers, (E) pentamers, (F) hexamers of WT Aβ 1–40 (upper panels), NSCR Aβ 1–40 (middle panels) and SCR Aβ 1–40 (lower panels), extracted from IM-MS data, with the height of each signal equal to the maximum value of the signal in the corresponding charge envelope. In SCR, DIM 7+ was missing, DIM 6+ was less intense, and DIM 4+ was more intense. The charge state envelope was in some instances (dimers, trimers, pentamers) shifted towards a lower charge in SCR. For trimers, the dominant form was 5+ in SCR and 7+ in both WT and NSCR, while the 8+ form was missing in SCR. TET 8+ and TET 9+ were missing in SCR but the remaining 7+, 6+, and 5+ signals retained the same mutual amplitude. A major PEN 9+ signal in WT and NSCR was missing in SCR, for which PEN 7+ was predominant. Hexameric signals are absent in SCR.

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques:

Journal: PLoS ONE

Article Title: Side-chain moieties from the N-terminal region of Aβ are Involved in an oligomer-stabilizing network of interactions

doi: 10.1371/journal.pone.0201761

Figure Lengend Snippet: For each selected m/z region, a profile of molecular species present at different drift times in the IM T-wave is shown, with each species assigned to a particular oligomeric charge state. Some profiles contain a single peak, indicating the presence of one form of a single collisional cross section, whereas other signals are split in the domain of the drift time into multiple species. (A) DIM 5+ —the dominant, extended form becomes more similar to the compact form in SCR. For TET 7+ (D) and TET 6+ (E), alternative structural forms of WT Aβ collapse in SCR to a single major form with intermediate drift time (and thus intermediate Ω), while TRI 7+ in SCR (B) exhibits a single form with the drift time closer to the compact form in WT. The split TRI 6+ signals as well as the TET 8+ signal (C), which are clear in WT and NSCR, become very weak in SCR. Signals were assigned to species as described before in Ref .

Article Snippet: The vectors of Aβ 1–40 scrambled sequence (SCR) and Aβ 1–40 scrambled sequence 1–16 (NSCR) in pET-30a(+) were purchased from GenScript and the peptides were expressed in Escherichia coli and then purified by HPLC as described previously [ ].

Techniques: