Fig 1.
The influence of: (a) temperature, (b) hydrolysis time, and (c) enzyme concentration on the DH (●) and LI (●) of the DLSH. Each data value is presented in the form of mean ± SE and the tests were conducted in triplicate.
Table 1.
The 20 experimental runs of the independent variables (X1, X2 and X3) and experimental values of responses which included DH (Y1) and IC50 of LI (Y2).
Each data values are presented in the form of mean ± SE and the tests were conducted in triplicate.
Table 2.
ANOVA for DH (Y1) and IC50 of LI (Y2).
Fig 2.
3D response surface plots and 2D contour plots for DH with the various factors: (a) The combined influence of temperature and time; (b) The combined influence of temperature and enzyme concentration, and (c) The combined influence enzyme concentration and time.
Fig 3.
3D response surface plots and 2D contour plots for LI with the various factors: (a) The combined influence of temperature and time; (b) The combined influence of temperature and enzyme concentration, and (c) The combined influence of time and enzyme concentration.
Table 3.
LI (IC50) results of DLSH ultrafiltered fractions.
Fig 4.
The RP-HPLC chromatogram for fraction <0.65 kDa of the original DLSH ultrafiltered fraction.
Table 4.
LI results of RP-HPLC fraction of DLSH.
Fig 5.
Amino acid sequence mass fragmentation spectrum and analysis for HPLC fraction F1 (GRSPDTHSG) by LC-Q-TOF-MS/MS.
Table 5.
The GRSPDTHSG peptide properties profiles.
Fig 6.
(a) The Lineweaver-Burk plot showing lipase inhibition with the GRSPDTHSG peptide both present and absent. 1/V and 1/S are the respective reciprocals of the velocity and substrate. Each of the points is presented in the form of mean value ± SE. Deviation with tests performed in triplicate, and (b) the Dixon plots showed the determination of the inhibitor constant (Ki) in the case of the non-competitive inhibition by GRSPDTHSG.
Table 6.
Kinetic parameters of lipase activity at varying GRSPDTHSG peptide concentrations.
Fig 7.
Molecular interaction between the GRSPDTHSG peptide and the porcine pancreatic lipase-colipase complex (1ETH).
(a) Broad perspective for the 3D illustration of the interactions between peptide-lipase complex. (b) 2D diagram of the anticipating interaction.
Table 7.
Intermolecular interactions between the GRSPDTHSG peptide and lipase residues.
Fig 8.
3T3-L1 cell viability after treatment for 72 h with different concentrations of (a) GRSPDTHSG, and (b) simvastatin. The graph presents the mean ± SE (n = 3). “ns” indicates not significance, while “*” indicates a significant difference compared to the untreated control (p < 0.001).
Fig 9.
Microscopic image of 3T3-L1 cells stained with Oil Red O (at 200 × magnification).
Fig 10.
The influence of the GRSPDTHSG peptide on adipogenesis of primary white adipocytes: (a) the investigated protein expression levels of PPAR-γ, SREBP-1c, and AMPK-α. C: undifferentiated cells; M: differentiated cells mode; S: simvastatin 10 μM, GRSPDTHSG peptide concentration at 0.25 (P1), 0.5 (P2) and 1.0 (P3) mM. (b) Detected bands of adipogenic factors. Results are presented after normalizing the values to β-actin level. The results are presented in the form of mean ± SE and the superscripts a-d on means represent significant difference (p < 0.05). Differentiated primary white adipocytes treated with the differentiation cocktail served as the positive control.
Fig 11.
The hypothesis proposes that the GRSPDTHSG peptide has the potential to diminish lipid components and inhibits the process of adipogenesis in 3T3-L1 cells.
AMPK: adenosine monophosphate-activated protein kinase; PPAR-γ: peroxisome proliferator-activated receptor gamma; SREBP-1c: sterol regulatory element-binding protein-1c. Biorender (©BioRender–biorender.com) was used to create the figure.