METHODS: ACE inhibitory proteins were isolated from P. cystidiosus based on the bioassay guided purification steps, i.e. ammonium sulphate precipitation, reverse phase high performance liquid chromatography and size exclusion chromatography. Active fraction was then analysed by LC-MS/MS and potential ACE inhibitory peptides identified were chemically synthesized. Effect of in vitro gastrointestinal digestions on the ACE inhibitory activity of the peptides and their inhibition patterns were evaluated.
RESULTS: Two potential ACE inhibitory peptides, AHEPVK and GPSMR were identified from P. cystidiosus with molecular masses of 679.53 and 546.36 Da, respectively. Both peptides exhibited potentially high ACE inhibitory activity with IC50 values of 62.8 and 277.5 μM, respectively. SEC chromatograms and BIOPEP analysis of these peptides revealed that the peptide sequence of the hexapeptide, AHEPVK, was stable throughout gastrointestinal digestion. The pentapeptide, GPSMR, was hydrolysed after digestion and it was predicted to release a dipeptide ACE inhibitor, GP, from its precursor. The Lineweaver-Burk plot of AHEPVK showed that this potent and stable ACE inhibitor has a competitive inhibitory effect against ACE.
CONCLUSION: The present study indicated that the peptides from P. cystidiosus could be potential ACE inhibitors. Although these peptides had lower ACE inhibitory activity compared to commercial antihypertensive drugs, they are derived from mushroom which could be easily obtained and should have no side effects. Further in vivo studies can be carried out to reveal the clear mechanism of ACE inhibition by these peptides.
OBJECTIVE: The objective of this study was to investigate the effects of four different polyols, namely, ethylene glycol, erythritol, xylitol and sorbitol on the acid-denatured states of CGB lectin.
METHODS: CGB lectin was subjected to acid denaturation at pH 2.5 and pH 1.5, both in the absence and presence of 30% (w/v) polyols, i.e. ethylene glycol, erythritol, xylitol and sorbitol. Thermal denaturation of the acid-denatured states was also studied in the absence and presence of these polyols. Different spectroscopic probes such as tryptophan fluorescence, ANS fluorescence and far-UV CD spectral signal were used to monitor structural changes in the acid-denatured states of CGB lectin in the presence of polyols.
RESULTS: Presence of erythritol, xylitol and sorbitol in the incubation mixture was found to stabilize the lectin at both pH 2.5 and pH 1.5, as evident from the burial of the hydrophobic clusters and decreased polarity around Trp residues. These polyols also stabilized the acid-denatured states of CGB lectin against thermal denaturation by shifting the thermal transition curves towards higher temperatures. Exposure of the acid-denatured states of CGB lectin, obtained at pH 2.5 and pH 1.5 to 61°C and 51°C, respectively, induced formation of non-native β-structures, compared to that present at 25°C, and this phenomenon was significantly suppressed in the presence of these polyols. Based on the spectral data, both sorbitol and erythritol appeared to exude better stabilizing effect. On the other hand, ethylene glycol was shown to destabilize the aciddenatured states of CGB lectin.
CONCLUSION: Thermal stabilization of the lectin was noticed in the presence of erythritol, xylitol and sorbitol at both pH 2.5 and pH 1.5. These polyols also stabilize the secondary and tertiary structures of the acid-denatured CGB lectin at 25°C. Ethylene glycol was proved to be a destabilizer of the acid-denatured CGB lectin.
METHOD: Literature search was performed within the PubMed, ScienceDirect.com and Google Scholar.
RESULTS: The presence of proline at the C-terminal tripeptide of ACE inhibitor can competitively inhibit the ACE activity. The effects of other amino acids are less studied leading to difficulties in predicting potent peptide sequences. The broad specificity of the enzyme may be due to the dual active sites observed on the somatic ACE. The inhibitors may not necessarily competitively inhibit the enzyme which explains why some reported inhibitors do not have the common ACE inhibitor characteristics. Finally, the in vivo assay has to be carried out before the peptides as the antihypertensive agents can be claimed. The peptides must be absorbed into circulation without being degraded, which will affect their bioavailability and potency. Thus, peptides with strong in vitro IC50 values do not necessarily have the same effect in vivo and vice versa.
CONCLUSION: The relationship between peptide amino acid sequence and inhibitory activity, in vivo studies of the active peptides and bioavailability must be studied before the peptides as antihypertensive agents can be claimed.