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The tenderization effect of a new elastase from Bacillus sp. EL31410 was investigated on beef meat. Meat tenderization was done by dipping the meat cut in a solution containing proteolytic enzymes after freeze-dehydration. It was found that a marination time of 4 h was enough for enzyme adsorption. The samples were treated for 4 h in different enzyme solutions and then was stored at 4 °C for 24, 48, 72 h, and subjected to texture measurement, sensory evaluations, biochemical analysis and histological observations. A marked decrease in hardness, by texture measurements, was observed in the meats with papain and elastase and higher sensory scores for tenderness were observed in the meats treated with enzymes than in the control. The papain-treated beef meat received the highest score for tenderness, but the scores given for juiciness and taste were lower than that of the control. Rapid increases of fragmentation of myofibrils from the enzyme-treated meat were observed in the first 24 h of storage, especially for papain-treated meat. Meantime, elastin of myofibrilar structure was selectively degraded by elastase compared with the control when stored at 4 °C for 48 h as shown by electron microscopy. These findings suggest that Bacillus elastase (EL31410) is a promising substitute for papain as a favourable meat tenderizer.