Zhiyong Li's research while affiliated with Soochow University (PRC) and other places

In this study, Ulva prolifera protein was used for preparing angiotensin-I converting enzyme (ACE)-inhibitory peptide via virtual gastrointestinal digestion and in silico screening. Some parameters of the obtained peptide, such as inhibition kinetics, docking mechanism, stability, transport pathway, were explored by Lineweaver-Burk plots, molecular docking, in vitro stimulate gastrointestinal (GI) digestion and Caco-2 cells monolayer model, respectively. Then, a novel anti-ACE peptide LDF (IC50, (1.66 ± 0.34) µmol/L) was screened and synthesized by chemical synthesis. It was a no-competitive inhibitor and its anti-ACE inhibitory effect mainly attributable to four Conventional Hydrogen Bonds and Zn701 interactions. It could keep activity during simulated GI digestion in vitro and was transported by peptide transporter PepT1 and passive-mediated mode. Besides, it could activate Endothelial nitric oxide synthase (eNOS) activity to promote the production of NO and reduce Endothelin-1 (ET-1) secretion induced by Angiotensin II (Ang II) in Human Umbilical Vein Endothelial Cells (HUVECs). Meanwhile, it could promote mice splenocytes proliferation in a concentration-dependent manner. Our study indicated that this peptide was a potential ingredient functioning on vasodilation and enhancing immunity.

The main protease (Mpro) is essential for the replication of SARS-COV-2 and therefore represents a promising anti-viral target. In this study, we screened Mpro inhibitory peptides from Ulva prolifera protein on in-silico proteolysis. Cytotoxicity analysis using the online toxic prediction tool ToxinPred revealed that all the peptides were non-cytotoxic. The hexapeptide (SSGFID) exhibited high Mpro inhibitory activity in molecular docking and its IC50 value was 139.40±0.82 µmol/L in vitro according to fluorescence resonance energy transfer assay (FRET). Quantitative real-time (qRT-) PCR results show that SSGFID could stimulate the expression of mitosis-related factors, including nuclear factor-κB, cyclin D1, and cyclin-dependent kinase 4, to promote the proliferation of mice splenocytes. Stability study revealed that SSGFID showed resistance against pepsin and trypsin but lost D (Asp) after pretreatment at 121 °C for 15 min. Besides, SSGFID was mainly transported through the Caco-2 cell monolayer by the peptide transporter PepT1 and passive-mediated transport during the transport study. Unfortunately, the peptide was also degraded by Caco-2 intracellular enzymes, and the transfer rate of intact peptide was 4.2%. Furthermore, Lineweaver–Burk plots demonstrated that SSGFID possessed a mixed inhibitory characteristic with Mpro. Our study indicated the potential of Ulva prolifera as antiviral and immune-enhancing functional food ingredients and nutraceuticals.

Ulva prolifera is the dominant species of “green tide”, and has higher tolerance to environmental stresses such as temperature. However, the molecular mechanisms are still unclear. Here, transcriptome analysis, Western blot and RT-qPCR analysis of U. prolifera suggested that, under temperature stresses (4°C, 36°C), the expression of PCNA and CyclinA was promoted, and the MAPK signaling was activated. Besides, the results showed that PCNA interacted with CyclinA. Interestingly, the expression of miR-2916 , which was predicted to bind PCNA at -552∼-772, was negatively correlated with the expression of PCNA under temperature stresses (4°C, 36°C). In addition, the results showed that low temperature (4°C) had no obvious effect on the survival, the formation of cell walls, and the division of protoplasts. However, high temperature (36°C) had obvious effect on them. PCNA inhibitors increased the sensitivity of the protoplasts under temperature stresses. Together, our results suggested PCNA regulating the proliferation in response to the temperature stress of U. prolifera was associated with miR-2916/PCNA/CyclinA/MAPK pathway. In conclusion, the study preliminarily illuminates the molecular mechanism in response to temperature stress of U. prolifera , and may provide a new insight for prevention of green tide.

In the present study, Ulva prolifera, an edible alga, was used to prepare angiotensin-I converting enzyme (ACE) inhibitory peptide. The algae protein was isolated and later hydrolyzed by five commercial enzymes (alcalase, papain, pepsin, trypsin, neutral protease), either individually or in combination. Hydrolysate, with the highest in vitro ACE inhibitory activity, was processed using the Sephadex-G100, ultrafiltration, HPLC-Q-TOF-MS, ADMET screening and molecular docking, respectively. The ACE inhibitory peptide DIGGL with a IC50 value of 10.32 ± 0.96 μM was then identified. The peptide against ACE by a non-competitive mode and mainly attributable to the three Conventional Hydrogen Bonds. It could activate Endothelial nitric oxide synthase activity in NO generation and reduce Endothelin-1 secretion induced by Angiotensin II in Human umbilical vein endothelial cells. Meanwhile, DIGGL could promote mice splenocytes proliferation, which was also effective when co-incubated with Con A or LPS, respectively. Besides, the anti-ACE peptide could remain active during the digestion of gastrointestinal proteases (pepsin-trypsin) in vitro.