The intermediate of Cys250-AdoHcy analyzed by MALDI-TOF-MS.

<p>(A) in GSH reaction system. (B) in AdoMet control. (C) in hAS3MT. (D) in reduced hAS3MT. (E) MS/MS spectra for the sequence at 1855.13 (m/z) of hAS3MT in GSH reaction system. Labels a, b, c, x, y, and z represent the cleavage manners of peptide. AdoMet control: 1.0 mM AdoMet was incubated with 2.0 µM hAS3MT at 37°C for 60 min.</p

Sequences for the peptide fragments of reduced hAS3MT analyzed by MALDI-TOF-MS.

<p>Values of observed M+H⁺ (Da) are the results of MALDI-TOF-MS in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110924#pone-0110924-g001" target="_blank">Figure 1b</a>.</p><p>Sequences for the peptide fragments of reduced hAS3MT analyzed by MALDI-TOF-MS.</p

Sequences for the AdoHcy-Cys250 peptide fragments at 1855.13 (m/z) analyzed by MALDI-TOF-MS/MS.

<p>Values of observed M+H⁺ (Da) are the results of MALDI-TOF-MS/MS in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110924#pone-0110924-g005" target="_blank">Figure 5e</a>. Mass for each fragments were calculated according to the standard fragmentation pattern.</p><p>Sequences for the AdoHcy-Cys250 peptide fragments at 1855.13 (m/z) analyzed by MALDI-TOF-MS/MS.</p

MALDI-TOF-MS spectra of the tryptic digests of C360S (A) and its reduced form (B).

<p>MALDI-TOF-MS spectra of the tryptic digests of C360S (A) and its reduced form (B).</p

Three-dimensional fluorescence spectra of (A) hAS3MT and (B) C250S.

<p>Each experiment was carried out three times.</p

MALDI-TOF-MS spectra of the tryptic digests of hAS3MT (A) and its reduced form (B). (C) MS/MS spectra for the sequence at 3451.17 (m/z).

<p>Labels a, b, c, x, y, and z represent the cleavage manners of peptides.</p

The effects of (A) GSH, (B) Cys, (C) DTT, and (D) TCEP on the Cys250-Cys32 disulfide bond.

<p>Before analysis, hAS3MT was respectively reduced by GSH (7 mM), Cys (10 mM), DTT (1.5 mM) or TCEP (0.7 mM) for 60 min at 37°C.</p

Fluorescence quenching of hAS3MT/C250S with AdoMet.

<p>(A, B) Stern-Volmer curves and (C, D) Double logarithm curves for the fluorescence quenching of hAS3MT and C250S with AdoMet at 350 nm, Ex: 276 nm. Solid line and dot line respectively represent the temperature at 27°C and 37°C. Each experiment was carried out three times.</p

MALDI-TOF-MS spectra for the digested hAS3MT after it catalyzed the iAs³⁺ methylation in GSH (A), Cys (B), DTT (C), or TCEP (D) reaction system.

<p>Before analysis, hAS3MT catalyzed the iAs³⁺ methyation in GSH, Cys, DTT, or TCEP reaction system at 37°C for 60 min.</p

Recovery effects of TCEP and DTT on the enzymatic methylation of iAs³⁺.

<p>(A) HPLC-ICP-MS spectra of arsenic species on hAS3MT. Control: hAS3MT (2.0 µM) catalyzed the iAs³⁺ methylation in the GSH reaction system for 60 min and dialysis against PBS. Dialysis-PBS, Dialysis-DTT (0.5 mM), and Dialysis-TCEP (0.7 mM): PBS, DTT, or TCEP separately incubated with the dialyzed hAS3MT in the presence of AdoMet (0.1 mM) at 37°C for another 60 min. (B) CD spectra and (C) corresponding secondary structure of hAS3MT. Enzyme: hAS3MT (2.0 µM) in PBS. <i>Error bars</i> represent S.D. from the mean of three independent experiments.</p