Proteases were isolated from leaf, bark and root of Choerospondias axillaris, locally called Lapsi. Choerospondias axillaris is a dioecous, deciduous fruit bearing plant with multiple daily uses. The protease was extracted with 0.1 M phosphate buffer of pH 7 and then precipitated successively with TCA and ammonium sulfate. The protease from leaf showed maximum activity at pH 9, temperature at 20°C. While, bark protease showed optimum pH 5 and temperature 60°C. In case of root the optimum pH was 10 and the optimum temperature 35°C. The optimum time of incubation leaf, bark and root was 15 minute. The bell shaped curve was obtained for the effect of enzyme conc. with optimum enzyme conc. 50 µg for leaf, 30 µg for bark and 50 µg for root. The K m and V max value of leaf were 5.61 µM and 185.18 pmol/ min, K m and V max value of bark were 2.36 µM and 82.64 pmol/min. While for root, the K m and V max values were 1.53 µM and 52.91 pmol/min.

The hypocholesterolemic peptide, LPYPR, was successfully introduced into the VR-1, VR-2, and VR-5 regions of the mung bean 8Sα globulin. The mutant protein (MP) has 96.69% structural homology and 97% sequence homology compared to the wild type (WT). Expression of the mutant protein in E. coli HMS174(DE3) was 40.66%, which was 144.42% higher than that of the WT. The WT protein and MP had MWs of about 48.4 and 48.7 kDa, respectively. These were purified using HIC and digested with trypsin. UPLC analysis of the tryptic digests of the MP revealed the successful release of the LPYPR peptide. Unlike the WT protein, cholesterol-binding capacity (mg/g sample) of the MP increased over time of tryptic digestion (average growth rate of 9.5% for crude MP and 12.5% for HIC-purified MP) for its undigested form (crude: 220.96 ± 8.65, purified: 214.71 ± 11.91), with maximum values of 380.76 ± 6.61 and 434.44 ± 10.88 were obtained for the 24-h digests of the crude and purified proteins, respectively. Similarly, the sodium taurocholate binding capacity (%) was also found to increase over time of tryptic digestion (average growth rate of 4% for crude MP and 5.67% for HIC-purified MP) for the tryptic digests of the MP. Minimum values for % bound sodium taurocholate was obtained with the undigested samples (crude: 46.71 ± 0.42, purified: 44.49 ± 0.13), while maximum values thereof were obtained with the 24-h digest samples (crude: 59.75 ± 0.30, purified 61.95 ± 0.51).