Yilmaz Halil, Yaradir Emine, Tunca Sedef
Faculty of Science, Molecular Biology and Genetics Department, Gebze Technical University, Gebze, 41400, Kocaeli, Türkiye.
Curr Microbiol. 2024 Dec 17;82(1):43. doi: 10.1007/s00284-024-04021-z.
The genus Streptomyces is a group of gram-positive bacteria that exhibit a distinctive growth pattern characterised by elongated, branched hyphae. Streptomyces coelicolor A3(2), which produces at least five different antibiotics, is a model organism that is widely used in genetic studies. There are very few studies in Streptomyces on the ATP-dependent Lon protease, which has very important functions in every organism and is particularly responsible for protein homeostasis. The aim of this study was to construct and characterize a recombinant S. coelicolor strain expressing the lon gene on a multicopy plasmid. For this purpose, the lon gene was first cloned in Escherichia coli under the control of the glycerol-inducible promoter of pSPG, and its expression in S. coelicolor A3(2) cells was demonstrated by RT-qPCR. In contrast with the initial hypothesis, increased lon expression did not affect cell growth seriously. Instead, it increased the cell's tolerance to osmotic and UV stress and led to a significant increase in antibiotic production. The recombinant strain produced 27 times more actinorhodin and 43 times more undecylprodigiosin than the wild-type strain after 120 h of fermentation. To our knowledge, this is the first study to demonstrate the effects of expression of the lon gene on a high copy number plasmid in Streptomyces.
链霉菌属是一组革兰氏阳性细菌,呈现出独特的生长模式,其特征为菌丝细长且有分支。天蓝色链霉菌A3(2)能产生至少五种不同的抗生素,是一种在遗传学研究中广泛使用的模式生物。在链霉菌中,关于ATP依赖型Lon蛋白酶的研究非常少,该蛋白酶在每种生物体中都具有非常重要的功能,尤其负责蛋白质稳态。本研究的目的是构建并表征在多拷贝质粒上表达lon基因的重组天蓝色链霉菌菌株。为此,首先将lon基因克隆到大肠杆菌中,置于pSPG的甘油诱导型启动子控制下,并通过RT-qPCR证明其在天蓝色链霉菌A3(2)细胞中的表达。与最初的假设相反,lon表达的增加并没有严重影响细胞生长。相反,它提高了细胞对渗透压和紫外线胁迫的耐受性,并导致抗生素产量显著增加。发酵120小时后,重组菌株产生的放线紫红素比野生型菌株多27倍,十一烷基灵菌红素多43倍。据我们所知,这是第一项证明lon基因在链霉菌中高拷贝数质粒上表达的影响的研究。