Discovery of a novel nanomolar angiotensin-I converting enzyme inhibitory peptide with unusual binding mechanisms derived from Chlorella pyrenoidosa.

Chlorella pyrenoidosa (C. pyrenoidosa) has been cultivated in large quantities, and was proven to be antihypertensive when consumed orally. However, the antihypertensive peptides derived from C. pyrenoidosa remains scarce. In this study, trypsin was chosen to prepare the hydrolysate of C. pyrenoidosa, which was then fractionated by column chromatography. And ninety-nine peptides were identified by LC-MS/MS, after which 10 peptides were chosen by docking-based virtual screening and demonstrated the ability to inhibit ACE. Among them, LVAKA (LV-5) had the lowest IC50 (26.66 µM). LV-5, LKKAP, and PGLRP were identified as non-competitive ACE inhibitory peptides that exhibited significant stability in the face of extreme pH and high temperatures. Insilico and in-vitro simulated gastrointestinal digestion revealed that these three peptides could release ACE inhibitory peptide fragments after digestion. The sequence optimization of LV-5 led to the identification of LRAKA (LR-5), which was recognized as a novel nanomolar ACE peptide with an IC50 of 350 nM in-vitro and a potent antihypertensive effect in-vivo. Moreover, molecular dynamic simulation indicated that LR-5 interacted with an unconventional binding site in ACE. These findings underscore the potential of Chlorella as a source of antihypertensive peptides and suggest a promising future for the use of Chlorella-derived peptides in the management of hypertension.

Inhibitory activity of a sulfated oligo-porphyran from Pyropia yezoensis against SARS-CoV-2.

COVID-19 caused by SARS-CoV-2 has spread around the world at an unprecedented rate. A more homogeneous oligo-porphyran with mean molecular weight of 2.1 kD, named OP145, was separated from Pyropia yezoensis. NMR analysis showed OP145 was mainly composed of →3)-ß-d-Gal-(1 â†’ 4)-α-l-Gal (6S) repeating units with few replacement of 3,6-anhydride, and the molar ratio was 1:0.85:0.11. MALDI-TOF MS revealed OP145 contained mainly tetrasulfate-oligogalactan with Dp range from 4 to 10 and with no more than two 3,6-anhydro-α-l-Gal replacement. The inhibitory activity of OP145 against SARS-CoV-2 was investigated in vitro and in silico. OP145 could bind to Spike glycoprotein (S-protein) through SPR result, and pseudovirus tests confirmed that OP145 could inhibite the infection with an EC50 of 37.52 µg/mL. Molecular docking simulated the interaction between the main component of OP145 and S-protein. All the results indicated that OP145 had the potency to treat and prevent COVID-19.

Fucoidan, as an immunostimulator promotes M1 macrophage differentiation and enhances the chemotherapeutic sensitivity of capecitabine in colon cancer.

Chemotherapy resistance is one of the most critical challenges in colorectal cancer (CRC) treatment. The occurrence and development of chemotherapy resistance closely related to the tumor immune microenvironment (TIME). As the most important immunosuppressive immune cells infiltrating into the TIME, macrophages are essential for chemotherapy resistance in CRC treatment. In this study, we found that a kind of fucoidan (FPS1M) induced macrophages differentiation to the M1 phenotype, and this transformation promoted cancer cells apoptosis both in vitro and in vivo. TNFα is a key mediator of FPS1M-induced tumorcidal activity of macrophages. Mechanistically, as a stimulator of TLR4, FPS1M enhanced macrophages glycolysis and regulated macrophages differentiation to the M1 phenotype by the activation of TLR4 mediated PI3K/AKT/mTOR signaling axis. In addition, FPS1M improved the immunosuppressed tumor microenvironment by increasing the infiltration of M1 macrophages in tumor tissue, which was conducive to improving the sensitivity of tumor to chemotherapy. Collectively, our findings demonstrated that FPS1M has the great potential to be used in tumor immunotherapy. The results also suggested that the combination of FPS1M with capecitabine is an alternative therapy method for colon cancer.

Isolation, identification and in vivo antihypertensive effect of novel angiotensin I-converting enzyme (ACE) inhibitory peptides from Spirulina protein hydrolysate.

The side effects of traditional antihypertensive drugs have driven people's interest in the discovery of novel angiotensin-I converting enzyme (ACE) inhibitory peptides with efficiency and safety. Spirulina possesses abundant proteins and is considered as a good source of bioactive peptides. To identify ACE inhibitory peptides, Spirulina protein was hydrolyzed by protease K, and the hydrolysate was separated by gel permeation chromatography and reverse phase HPLC. Thirty peptide sequences were identified from the ACE inhibitory fractions by LC-MS/MS, and 15 potential ACE inhibitory peptides were further selected by combined virtual screening and experimental investigation. Among the identified Spirulina peptides, TVLYEH (SpH-6, IC50 = 2.88 µM) and LQAGGLF (SpH-7, IC50 = 66.83 µM) were found to possess potent ACE inhibitory activity. The inhibition patterns of SpH-6 and SpH-7 were characterized as competitive inhibition. Molecular docking indicated that SpH-6 and SpH-7 could bind to the active pockets of ACE by forming hydrogen bonds, salt bridges and pi-pi stacking. Both SpH-6 and SpH-7 exhibited stable ACE inhibitory activity under different pH (2, 4, 6, 8, 10, 12) and temperature (0, 20, 40, 60, 80, 100 °C) conditions. The ACE inhibitory activity of SpH-6 decreased slightly after simulated gastrointestinal digestion, and SpH-7 was unstable against digestive enzymes. The in vivo antihypertensive effect demonstrated that oral administration of SpH-6 significantly reduced the systolic blood pressure and diastolic blood pressure of spontaneously hypertensive rats. The present study revealed the potential antihypertensive effects of SpH-6 and indicated that SpH-6 and Spirulina protein hydrolysate could be applied as nutritional supplements for blood pressure control.

Discovery of a novel jellyfish venom metalloproteinase inhibitor from secondary metabolites isolated from jellyfish-derived fungus Aspergillus versicolor SmT07.

The major jellyfish stings that occur in China are caused by scyphozoan Nemopilema nomurai, whose venom exhibits significant metalloproteinase activity that contributes to the toxic effects of jellyfish envenomation. Researching effective inhibitors suppressing the metalloproteinase activity of jellyfish venom represents a new attempt to cure jellyfish envenomations. In the present study, secondary metabolites produced by the jellyfish-associated fungus Aspergillus versicolor SmT07 were isolated and evaluated for their anti-proteolytic activities. Two xanthones, sterigmatocystin (JC-01) and oxisterigmatocystin C (JC-06), and four alkaloids, cottoquinazoline A (JC-02), phenazine-1-carboxylic acid (JC-03), viridicatin (JC-04) and viridicatol (JC-05), were isolated and identified. Only phenazine-1-carboxylic acid (PCA) showed significant anti-proteolytic activity of jellyfish venom assayed on azocasein, and the IC50 value was 2.16 mM. PCA also significantly inhibited fibrinogenolytic activity, protecting the Bß chain of fibrinogen from degradation when preincubated with jellyfish venom at a ratio of >1:0.6 (PCA:venom, w/w). Molecular docking with several well-characterized snake venom metalloproteinases suggested the venom metalloproteinases inhibitory property of PCA by forming complex interactions with the active site via hydrogen bonds, π-π stacking and salt bridges, which was distinct from the binding mode of batimastat. The present study represents the first study identifying natural jellyfish venom metalloproteinase inhibitors from marine natural products, which may provide an alternative to develop therapeutic agents for treating jellyfish envenomations.