ACE Inhibitory Activity and Molecular Docking of Gac Seed Protein Hydrolysate Purified by HILIC and RP-HPLC

Abstract

Gac (Momordica cochinchinensis Spreng.) seed proteins (GSPs) hydrolysate was investigated for angiotensin I-converting enzyme (ACE) inhibitory activities. GSPs were hydrolyzed under simulated gastrointestinal digestion using a combination of enzymes, including pepsin, trypsin, and chymotrypsin. The screening of ACE inhibitory peptides from GSPs hydrolysate was performed using two sequential bioassay-guided fractionations, namely hydrophilic interaction liquid chromatography (HILIC) and reversed-phase high-performance liquid chromatography (RP-HPLC). Then, the peptides in the fraction with the highest ACE inhibitory activity were identified by LC-MS/MS. The flow-through (FT) fraction showed the most potent ACE inhibitory activity when HILIC fractionation was performed. This fraction was further separated using RP-HPLC, and the result indicated that fraction 8 (RP-F8) showed the highest ACE inhibitory activity. In the HILIC-FT/RP-F8 fraction, 14 peptides were identified using LC-MS/MS analysis coupled with de novo sequencing. These amino acid chains had not been recorded previously and their ACE inhibitory activities were analyzed in silico using the BIOPEP database. One fragment with the amino acid sequence of ALVY showed a significant ACE inhibitory activity (7.03 ± 0.09 µM). The Lineweaver-Burk plot indicated that ALVY is a competitive inhibitor. The inhibition mechanism of ALVY against ACE was further rationalized through the molecular docking simulation, which revealed that the ACE inhibitory activities of ALVY is due to interaction with the S1 (Ala354, Tyr523) and the S2 (His353, His513) pockets of ACE. Bibliographic survey allowed the identification of similarities between peptides reported as in gac fruit and other proteins. These results suggest that gac seed proteins hydrolysate can be used as a potential nutraceutical with inhibitory activity against ACE.

A novel angiotensin-I-converting enzyme inhibitory peptide from oyster: Simulated gastro-intestinal digestion, molecular docking, inhibition kinetics and antihypertensive effects in rats

“…The inhibitory patterns of the ACEI peptides mainly contain competitive inhibition, non-competitive inhibition, and mixed inhibition according to their various structures. For example, competitive ACEI peptides, namely ALVY from Gac Seed Protein Hydrolysates, PRY derived from potato patatin, and SSYYPFK from naked oat globulin hydrolysates, have been published ( 41 , 42 ). Non-competitive ACE inhibitors, such as VGLPNSR and QAGLSPVR, have been reported from tilapia skin gelatin, AVKVL, YLVR, and TLVGR derived from hazelnut (Corylusheterophylla Fisch), and ETSGMKPTEL and ISSMGILVCL isolated from longan seeds ( 32 , 37 ).…”

Preparation and activity evaluation of angiotensin-I converting enzyme inhibitory peptides from protein hydrolysate of mulberry leaf

“…Molecular docking was performed between the identified peptides and the crystal structure of the human angiotensin-I converting enzyme (1O8A) according to the method of Samuchaya et al ( 29 ). The semi-flexible docking of 1O8A to ACE inhibitory peptides was calculated using molecular docking software MOE.…”

Identification of dipeptidyl peptidase IV inhibitory peptides derived from gac seed protein hydrolysate using hydrophilic interaction liquid chromatography and reversed-phase high-performance liquid chromatography

“…Gac seeds have been disregarded in many countries where this fruit is grown (Abdulqader et al, 2019). However, gac seeds have great potential to serve as raw material for medicine preparation, nutraceuticals and functional foods because these seeds contain abundant proteins and bioactive compounds (Ngamsuk et al, 2020). Plant proteins have been known to aid in the impediment of many diseases such as diabetes (Arnoldi et al, 2015).…”

“…Active protein hydrolysate is generally composed of peptides with diverse weights, molecular sizes, hydrophobicity and physicochemical properties, with fractionation and purification of the peptides achieved using multidimensional column chromatography, such as solid-phase extraction (SPE), strong cation exchange (SCX), size-exclusion chromatography (SEC), hydrophilic interaction liquid chromatography (HILIC), reversed-phase high-performance liquid chromatography (RP-HPLC) and ion-exchange chromatography (IEC; Liu et al, 2017;Liu et al, 2019;Ngamsuk et al, 2020;Sutopo et al, 2020). Sungthong and Gerdvutgoa (1999) reported that the extract from animal feed purified using SPE was purer in solution than from using other methods.…”