Effects of different buffers on the thermostability and autolysis of a cold-adapted protease MCP-01
Effects of different buffers on the thermostability and autolysis of a cold-adapted protease MCP-01
复制标题
DOI：
10.1023/a:1022425621742
复制
发表时间：
2002-11-01
期刊：
影响因子：
--
通讯作者：
Gao, PJ
中科院分区：
文献类型：
--
作者：
Chen, XL;Sun, CY;Gao, PJ
冷适应蛋白酶 MCP-01 是从深海耐冷细菌 Pseudoaltermonas sp. 中获得的。 SM9913。研究了浓度均为 50 mmol/l 的四种不同缓冲液对其热稳定性和自溶作用的影响。通过毛细管电泳研究了MCP-01的自溶过程。 MCP-01的热稳定性按以下顺序依次增加：碳酸盐、Tris、磷酸盐、硼酸盐。酪蛋白水解的最适温度也以相同的顺序增加。这表明MCP-01的构象是灵活的，其自溶敏感性受到缓冲液中的一些因素（例如电荷和离子种类）的影响。结果还表明，不同的缓冲液除了影响自溶速度外，还会产生不同的自溶产物模式。在碳酸盐缓冲液、Tris缓冲液、磷酸盐缓冲液和硼酸盐缓冲液中，MCP-01的自溶模式不同。这些结果表明，蛋白酶MCP-01在不同的缓冲液中可能具有不同的构象，从而在酶表面暴露出不同的自溶位点。此外，活性的丧失与四种不同缓冲液中的自溶速度相关，表明自溶可能是酶热稳定性低的原因。
A cold-adapted protease MCP-01 was obtained from deep-sea psychrotrophic bacterium Pseudoaltermonas sp. SM9913. The effects of four different buffers, all at 50 mmol/l concentration, on its thermostability and autolysis were studied. The autolysis process of MCP-01 was studied by capillary electrophoresis. The thermostability of MCP-01 increased successively in the following order: carbonate, Tris, phosphate, borate. The optimum temperature for casein hydrolysis also increased in the same order. This suggested that the conformation of MCP-01 was flexible and its autolytic susceptibility was affected by some factors in the buffers such as charge and ionic species. The results also showed that different buffers, in addition to affecting the autolysis speed, gave different patterns of autolysis products. In carbonate buffer, Tris buffer, phosphate buffer and borate buffer, the autolysis patterns of MCP-01 were different. These results suggested that protease MCP-01 probably have different conformations in different buffers, thus exposing different autolysis sites on the enzyme surface. In addition, the loss of activity correlated with the speed of autolysis in the four different buffers, showing that autolysis may be a reason for the low thermostability of the enzyme.