Abstract:To explore the related proteins secreted by Lactobacillus acidophilus and their functional roles in promoting gut homeostasis, the extracellular protein crude extract was obtained from L. acidophilus solid culture, and each fraction of the extracellular proteins was collected after purification and isolation using Sephadex G-100 gel chromatography. The amount of protein and molecular mass of each protein fraction were determined by Coomassie brilliant blue method and SDS-PAGE electrophoresis, respectively. Then, the inhibitory effect of each L. acidophilus extracellular protein fraction on the proliferation of HT-29 cells was investigated by 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay. Two protein peaks were obtained after purification, their molecular weights were 67 ku and 37 ku, respectively, and their protein contents were 0.066 mg/mL and 0.021 mg/mL, respectively. HT-29 cells were treated with both 67 ku and 37 ku extracellular proteins at concentrations of 0.001 μg/mL, 0.01 μg/mL, 0.1 μg/mL, and 1 μg/mL for 12 h, 24 h, 48 h, or 72 h, respectively. Both proteins showed an inhibitory effect on the proliferation of HT-29 cells in a time- and dose-dependent manner. Among the tested concentrations, the inhibition rates of the 67 ku and 37 ku extracellular proteins at μg/mL on HT-29 cells were (38.41±1.94)% and (45.06±1.58)%, respectively. In conclusion, L. acidophilus cultured on solid medium produced two types of extracellular proteins (37 ku and 67 ku), which inhibited the proliferation of HT-29 cells to different degrees, and the 37 ku extracellular protein exhibited a relatively strong inhibitory effect.
