克隆、鉴定和多个染色体整合一个芽孢杆菌碱性蛋白酶基因。
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van der拉恩说道JC, Gerritse G, Mulleners LJ, van der Hoek RA, Quax WJ
克隆、鉴定和多个染色体整合一个芽孢杆菌碱性蛋白酶基因。
:环境Microbiol。1991年4月;57 (4):901 - 9。
- PubMed ID
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2059048 (在PubMed]
- 文摘
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细胞外的芽孢杆菌蛋白酶在洗涤剂粉作为添加剂。我们发现产生的芽孢杆菌菌株与极碱性pH值最佳蛋白酶;这种蛋白酶适用于现代碱性洗涤剂粉末。alkalophilic株芽孢杆菌alcalophilus PB92基因编码这个高碱性丝氨酸蛋白酶是克隆和特征。序列分析显示一个开放阅读框的380个氨基酸组成的信号肽氨基酸(27),一个prosequence(84个氨基酸),和一个成熟的269个氨基酸的蛋白。氨基酸的比较与其他丝氨酸蛋白酶与蛋白酶YaB显示良好的同源性,这也是由一个alkalophilic杆菌菌株。都显示出温和与枯草杆菌蛋白酶同源,但显示一些显著的差异从中性杆菌产生的枯草杆菌蛋白酶。PB92蛋白酶的prosequence已经没有明显的同源性prosequences枯草杆菌蛋白酶。大量的负电荷氨基酸残基构成的prosequences PB92蛋白酶尤其显著。克隆的基因被用来提高蛋白酶的生产水平。 For this purpose the strategy of gene amplification in the original alkalophilic Bacillus strain was chosen. When introduced on a multicopy plasmid, the recombinant strain was unstable; under production conditions, plasmid segregation occurred. More stable ways of gene amplification were obtained by chromosomal integration. This was achieved by (i) homologous recombination, resulting in a strain with two tandemly arranged genes, and (ii) illegitimate recombination, resulting in a strain with a second copy of the protease gene on a locus not adjacent to the originally present gene. Both strains showed increased production and were more stable than the plasmid-containing strain. Absolute stability was only found when nontandem duplication occurred. This method of gene amplification circumvents stability problems often encountered in gene amplification in Bacillus species when plasmids or tandemly arranged genes in the chromosome are used.