Wittchen K D, Meinhardt F
Institut für Mikrobiologie, Westfälische Wilhelms-Universität, Münster, Germany.
Appl Microbiol Biotechnol. 1995 Mar;42(6):871-7. doi: 10.1007/BF00191184.
An efficient method for gene replacement in Bacillus megaterium was developed and used to inactivate the chromosomal neutral protease gene (nprM) from strain DSM319. A temperature-dependant suicide vector was constructed to allow replacement of the normal chromosomal copy with an altered version of the nprM gene. One mutant B. megaterium MS941 was selected for further characterization. Measurement of extracellular protease activity from strain MS941 indicated the existence of an additional minor extracellular protease in B. megaterium. Inhibitor studies revealed that this minor protease, comprising only 1.4% of the wild-type total extracellular protease activities, is a serine-type enzyme.
开发了一种在巨大芽孢杆菌中进行基因替换的有效方法，并用于使DSM319菌株的染色体中性蛋白酶基因（nprM）失活。构建了一种温度依赖性自杀载体，以便用nprM基因的改变版本替换正常的染色体拷贝。选择了一个突变型巨大芽孢杆菌MS941进行进一步表征。对MS941菌株细胞外蛋白酶活性的测量表明，巨大芽孢杆菌中存在另一种次要的细胞外蛋白酶。抑制剂研究表明，这种仅占野生型总细胞外蛋白酶活性1.4%的次要蛋白酶是一种丝氨酸型酶。
