Antitumor and HIV-1 Reverse Transcriptase Inhibitory Activities of a Hemagglutinin and a Protease Inhibitor from Mini-Black Soybean
Figure 5
(a) Determination of IC50 values of trypsin- and chymotrypsin-inhibitory activities of mini-black soybean protease inhibitor (MBSTI) and soybean trypsin inhibitor (STI). Results are means ± SD (). The protease inhibitor was incubated with bovine pancreatic trypsin and casein (for assay of trypsin inhibitory activity) or with N-α-benzoyl-L-tyrosyl ethyl ester hydrochloride (for assay of chymotrypsin inhibitory activity) and bovine pancreatic chymotrypsin for 15 minutes in 0.1 M Tris-HCl buffer (pH 7.4) before addition of 5% trichloroacetic acid. The reaction mixture was centrifuged and OD280 of the supernatant containing the tryptic fragments of casein was read. Background values were determined and subtracted before % inhibition values were calculated. Data points bearing the same letter represent statistically significant difference () when the data were analyzed by ANOVA followed by Duncan’s multiple range test. (b) Effect of dithiothreitol (DTT) on trypsin-inhibitory activity of 2.4 μM mini-black soybean (MBSPI) and 2.8 μM trypsin inhibitor from soybean (STI) after incubation at 37°C for 25 minutes. Results are means ± SD (). The IC50 of DTT was about 30 mM on MBSPI and lower than 10 mM on STI. The protease inhibitor (1.8 mM) was incubated with dithiothreitol at various concentrations before termination of reaction with iodoacetamide and determination of remaining trypsin-inhibitory activity. Background values were determined and subtracted before % inhibition values were calculated. Data points bearing the same letter represent statistically significant difference () when the data were analyzed by ANOVA followed by Duncan’s multiple range test.
