The regulatory mechanism of pulsed electric field (PEF) targeting at C-terminal glutamine of shrimp antioxidant peptide QMDDQ based on MD simulation
Introduction
PEF treatment have become an emerging technique compared to other non-thermal treatment technologies because it has incomparable advantages in many aspects (Fauster et al., 2020; Martín-García et al., 2020). It is noteworthy that PEF processing could improve the biological activity of food-borne substances. Zhang et al. investigated the effects of PEF treatment on the activity of enzyme, which was significantly (P < 0.05) increased and achieved the highest value at 12 kV/cm (F. Zhang, Tian, Du, & Fang, 2017). In addition to the above-mentioned biological activity, PEF processing can also be widely used to improve the antioxidant activity of peptides. Liang et al. explored the PEF processing on pine nut antioxidant peptides, which of the DPPH and ABTS radical inhibition capacities were increased at 2400 Hz and 15 kV/cm (R. Liang, Z. Zhang, & S. Lin, 2017a). Wang et al. reported that the DPPH radical inhibition activity of glutathione increased by 15.57% at 2549.08 Hz and 9.74 kV/cm (Wang et el., 2014) Nevertheless, there are few reports on the information about the relationship between the antioxidant activity and structure characterization of PEF-treated peptides. Therefore, it is continuously urgent to focus on PEF-treated peptides and further explore the correlation between antioxidant capacity and structure characterization.
Currently, the combination of nuclear magnetic resonance (NMR) and computational techniques is the most extensively used of small molecule conformation analysis technique (Nguyen, Schwochert, Tantillo, & Lokey, 2018). Molecule dynamics (MD) simulation is an efficient tool that can simulate the structure and dynamics in food-borne proteins after applying electric field stress (Vanga, Wang, Singh, & Raghavan, 2019). Through this we could not only get the atomic trajectory, but also get a variety of observation like to do the same experiment (Wang, Yan, Yao, & Li, 2008). Vanga, Singh, & Raghavan, investigated that the structure conformation of peanut allergen Ara h 6 changed after adding thermal and electric fields (Vanga et al.). However, the effect of PEF processing on biological activity and spatial conformation of peptides still lags behind the experimental study. Therefore, we could combine molecular dynamics simulation with structural characterization to explore the mechanism.
Previously, the sequence of Gln-Met-Asp-Asp-Gln (QMDDQ) was selected as the best antioxidant peptide among four similar amino acid sequences in terms of its good characteristics for the hydroxyl free radicals scavenging activity through EPR, the antioxidant ability (SOD, ROS and LDH) in PC12 cells, as well as the oxidative stress by flow cytometry and LSCM (Wu, Sun, Ding, Zhu, & Lin, 2019). In the current study, the effect of PEF processing on the antioxidant activity of the synthetic peptide (QMDDQ) was measured by DPPH radical and ABTS radical cation scavenging assays. Furthermore, the underlying mechanism of PEF-treated QMDDQ was identified by MIR and CD spectroscopy, and multiple interactions of residues in QMDDQ were further explored by using two-dimensional (2D) NMR and MD simulations. The purpose of this study was to illustrate the correlation between structure and function of the peptide by PEF and further determine the target sites.
Section snippets
Materials and reagents
The shrimp peptide QMDDQ was chemically synthesized by Hefei Saimanuo Biological Technology Co. (Hefei, Anhui Province, China). The molecular weight of the QMDDQ was 678.20 Da, and the purity was 99.40%. DPPH, ABTS, and dimethyl sulfoxide-D6 (DMSO-D6) were obtained from Sigma-Aldrich Co. (St Louis, MO, USA). Other reagents were obtained from Tianjin Damao chemical reagent factory (Tianjin, China) and were all analytical grade purity.
PEF treatment on QMDDQ
The self-designed PEF system was used to process peptide
Effects of PEF processing on antioxidant capacity of QMDDQ
In the present study, QMDDQ was treated by PEF at 4000 Hz with different electric field intensities (0, 10, 20, 30, 40 kV/cm). Fig. 1 showed the effects of different electric field intensities on antioxidant capacity of the peptides. From Fig. 1(a), the DPPH scavenging of QMDDQ was significantly (P < 0.05) increased and achieved maximum activity (47.07 ± 0.90%) at 40 kV/cm. Comparing with the untreated sample, the ABTS•+ scavenging activity of QMDDQ were significantly (P < 0.05) increased by
Conclusions
The aim of this study was to locate the PEF-induced target regulatory sites of the shrimp peptide QMDDQ. The DPPH and ABTS inhibition activities of the peptide were significantly (P < 0.05) increased after PEF treatment. The primary structure of the QMDDQ was not affected by PEF, whereas the β-sheet was converted into random coil in the peptide solution, thereby reducing the steric hindrance of the free radical. Moreover, NOESY spectrum further revealed that the change of antioxidant ability
CRediT authorship contribution statement
Shuyu Zhang: Writing - original draft, Writing - review & editing. Libo Qi: Investigation, Formal analysis. Dongmei Li: Resources, Validation, Investigation. Limin Zhong: Writing - review & editing. Dan Wu: Validation, Formal analysis. Songyi Lin: Conceptualization, Methodology, Supervision.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work.
Acknowledgements
The authors acknowledge the financial support provided by the National Key Research and Development Program of China (2017YFD0400500).
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