2 research outputs found
Improved N<sup>α</sup>‑Acetylated Peptide Enrichment Following Dimethyl Labeling and SCX
Protein
N-terminal acetylation is one of the most common modifications occurring
co- and post-translationally on either eukaryote or prokaryote proteins.
However, compared to other protein modifications, the physiological
role of protein N-terminal acetylation is relatively unclear. To explore
the biological functions of protein N-terminal acetylation, a robust
and large-scale method for qualitative and quantitative analysis of
this modification is required. Enrichment of N<sup>α</sup>-acetylated
peptides or depletion of the free N-terminal and internal tryptic
peptides prior to analysis by mass spectrometry are necessary based
on current technologies. This study demonstrated a simple strong cation
exchange (SCX) fractionation method to selectively enrich N<sup>α</sup>-acetylated tryptic peptides via dimethyl labeling without the need
for tedious protective labeling and depleting procedures. This method
was introduced for the comprehensive analysis of N-terminal acetylated
proteins from HepG2 cells. Several hundred N-terminal acetylation
sites were readily identified in a single SCX flow-through fraction.
Moreover, the N<sup>α</sup>-acetylated peptides of some protein
isoforms were simultaneously observed in the SCX flow-through fraction,
which indicated that this approach can be utilized to discriminate
protein isoforms with very similar full sequences but different N-terminal
sequences, such as β-actin/γ-actin, ERK1/ERK2, α-centractin/β-centractin,
and ADP/ATP translocase 2 and 3. Compared to other methods, this method
is relatively simple and can be directly implemented in a two-dimensional
separation (SCX-RP)-mass spectrometry scheme for quantitative N-terminal
proteomics using stable-isotope dimethyl labeling
Anti-α-glucosidase and Anti-dipeptidyl Peptidase-IV Activities of Extracts and Purified Compounds from Vitis thunbergii var. <i>taiwaniana</i>
Ethanol extracts (Et) from the stem
(S) and leaf (L) of Vitis thunbergii var. <i>taiwaniana</i> (VTT) were used to investigate
yeast α-glucosidase and porcine
kidney dipeptidyl peptidase-IV (DPP-IV) inhibitory activities. Both
VTT-Et showed complete α-glucosidase inhibition at 0.1 mg/mL;
VTT-S-Et and VTT-L-Et showed 26 and 11% DPP-IV inhibition, respectively,
at 0.5 mg/mL. The VTT-Et interventions (20 and 50 mg/kg) resulted
in improvements in impaired glucose tolerance of diet-induced obese
rats. (+)-Hopeaphenol, (+)-vitisin A, and (−)-vitisin B were
isolated from the ethyl acetate fractions of S-Et and showed yeast
α-glucosidase inhibition (IC<sub>50</sub> = 18.30, 1.22, and
1.02 μM) and porcine kidney DPP-IV inhibition (IC<sub>50</sub> = 401, 90.75, and 15.3 μM) compared to acarbose (6.39 mM)
and sitagliptin (47.35 nM), respectively. Both (+)-vitisin A and (−)-vitisin
B showed mixed noncompetitive inhibition against yeast α-glucosidase
and porcine kidney DPP-IV, respectively. These results proposed that
VTT extracts might through inhibitions against α-glucosidase
and DPP-IV improve the impaired glucose tolerance in diet-induced
obese rats