Wolański Marcin, Krawiec Michał, Schwarz Paul N, Stegmann Evi, Wohlleben Wolfgang, Buchmann Anina, Gross Harald, Eitel Michael, Koch Pierre, Botas Alma, Méndez Carmen, Núñez Luz Elena, Morís Francisco, Cortés Jesus, Zakrzewska-Czerwińska Jolanta
Faculty of Biotechnology University of Wrocław Wrocław Poland.
Department of Microbiology and Biotechnology Interfaculty Institute of Microbiology and Infection Medicine University of Tübingen Tübingen Germany.
Eng Life Sci. 2020 Nov 4;21(1-2):4-18. doi: 10.1002/elsc.202000038. eCollection 2021 Jan.
Brasilicardin A (BraA) is a promising immunosuppressive compound produced naturally by the pathogenic bacterium IFM 0406. Heterologous host expression of brasilicardin gene cluster showed to be efficient to bypass the safety issues, low production levels and lack of genetic tools related with the use of native producer. Further improvement of production yields requires better understanding of gene expression regulation within the BraA biosynthetic gene cluster (Bra-BGC); however, the only so far known regulator of this gene cluster is Bra12. In this study, we discovered the protein LysRNt, a novel member of the LysR-type transcriptional regulator family, as a regulator of the Bra-BGC. Using in vitro approaches, we identified the gene promoters which are controlled by LysRNt within the Bra-BGC. Corresponding genes encode enzymes involved in BraA biosynthesis as well as the key Bra-BGC regulator Bra12. Importantly, we provide in vivo evidence that LysRNt negatively affects production of brasilicardin congeners in the heterologous host . Finally, we demonstrate that some of the pathway related metabolites, and their chemical analogs, can interact with LysRNt which in turn affects its DNA-binding activity.
巴西卡地菌素A（BraA）是一种由致病性细菌IFM 0406天然产生的有前景的免疫抑制化合物。巴西卡地菌素基因簇的异源宿主表达被证明能有效绕过与使用天然生产者相关的安全问题、低产量水平和缺乏遗传工具等问题。进一步提高产量需要更好地理解巴西卡地菌素A生物合成基因簇（Bra-BGC）内的基因表达调控；然而，到目前为止该基因簇唯一已知的调控因子是Bra12。在本研究中，我们发现了LysRNt蛋白，它是LysR型转录调控因子家族的一个新成员，作为Bra-BGC的调控因子。使用体外方法，我们确定了Bra-BGC内受LysRNt控制的基因启动子。相应的基因编码参与巴西卡地菌素A生物合成的酶以及关键的Bra-BGC调控因子Bra12。重要的是，我们提供了体内证据，表明LysRNt对异源宿主中巴西卡地菌素类似物的产生有负面影响。最后，我们证明了一些与该途径相关的代谢物及其化学类似物可以与LysRNt相互作用，进而影响其DNA结合活性。
