Fuchs P G, Dobrzański W T
J Bacteriol. 1978 Oct;136(1):295-303. doi: 10.1128/jb.136.1.295-303.1978.
Trypsin and other proteolytic enzymes, added together with transforming DNA or during cell-DNA contact to competent cultures of several streptococcal strains, enchanced (10 to 600%) the yield of genetic transformation (stimulation). With few exceptions, the level of stimulation was high (over 100%) when competence was low (below 2%). Stimulation was caused by the action of an enzyme on competent cells and not on any other component of transformation mixture. The phenomenon occurred when the enzyme was added to the culture not earlier than 7 min before and not later than 5 min after the period of cell-DNA contact. The presence of trypsin during cell-DNA contact caused: (i) the alterations at cell surface, demonstrated by electron microscopy, increased release of 3H-amino acid-labeled material, and higher cell susceptibility to autolysis; (ii) the increase of both total and irreversible binding of DNA by the cells; and (iii) the decrease of early nucleolytic degradation of DNA by cells. These and other data point to the importance of a delicate balance of recipient cell's surface nuclease activities in the effectiveness of transformation process. It is also possible that trypsin eliminates an unknown cellular factor which obstructs DNA-cell receptors interaction.
将胰蛋白酶和其他蛋白水解酶与转化DNA一起添加,或在细胞与DNA接触期间添加到几种链球菌菌株的感受态培养物中,可提高(10%至600%)遗传转化的产量(刺激作用)。除少数例外,当感受态较低(低于2%)时,刺激水平较高(超过100%)。刺激是由酶对感受态细胞的作用引起的,而不是对转化混合物的任何其他成分的作用。当酶在细胞与DNA接触期之前不早于7分钟且不晚于5分钟添加到培养物中时,就会出现这种现象。细胞与DNA接触期间胰蛋白酶的存在导致:(i)通过电子显微镜观察到细胞表面的改变、3H-氨基酸标记物质释放增加以及细胞对自溶的敏感性更高;(ii)细胞对DNA的总结合和不可逆结合均增加;(iii)细胞对DNA的早期核酸降解减少。这些以及其他数据表明,受体细胞表面核酸酶活性的微妙平衡在转化过程的有效性中很重要。也有可能胰蛋白酶消除了一种阻碍DNA-细胞受体相互作用的未知细胞因子。