Rajkarnikar Arishma, Kwon Hyung-Jin, Suh Joo-Won
Department of Biological Science, Division of Bioscience and Bioinformatics, Myongji University, Yongin 449-728, Republic of Korea.
Biochem Biophys Res Commun. 2007 Nov 16;363(2):322-8. doi: 10.1016/j.bbrc.2007.08.169. Epub 2007 Sep 6.
Adenosine kinase (ADK) catalyses phosphorylation of adenosine (Ado) and generates adenosine monophosphate (AMP). ADK gene (adk(Sli), an ortholog of SCO2158) was disrupted in Streptomyces lividans by single crossover-mediated vector integration. The adk(Sli) disruption mutant (Deltaadk(Sli)) was devoid of sporulation and a plasmid copy of adk(Sli) restored sporulation ability in Deltaadk(Sli), thus indicating that loss of adk(Sli) abolishes sporulation in S. lividans. Ado supplementation strongly suppressed sporulation ability in S. lividans wild-type (wt), supporting that disruption of adk(Sli) resulted in Ado accumulation, which in turn suppressed sporulation. Cell-free experiments demonstrated that Deltaadk(Sli) lacked ADK activity and in vitro characterization confirms that adk(Sli) encodes ADK. The intracellular level of Ado was highly elevated while the AMP level was significantly reduced after loss of adk(Sli) while Deltaadk(Sli) displayed no significant derivation from wt in the levels of S-adenosylhomocysteine (SAH) and S-adenosylmethionine (SAM). Notably, Ado supplementation to wt lowered AMP content, albeit not to the level of Deltaadk(Sli), implying that the reduction of AMP level is partially forced by Ado accumulation in Deltaadk(Sli). In Deltaadk(Sli), actinorhodin (ACT) production was suppressed and undecylprodigiosin (RED) production was dramatically enhanced; however, Ado supplementation failed to exert this differential control. A promoter-probe assay verified repression of actII-orf4 and induction of redD in Deltaadk(Sli), substantiating that unknown metabolic shift(s) of ADK-deficiency evokes differential genetic control on secondary metabolism in S. lividans. The present study is the first report revealing the suppressive role of Ado in Streptomyces development and the differential regulatory function of ADK activity in Streptomyces secondary metabolism, although the underlying mechanism has yet to be elucidated.
腺苷激酶（ADK）催化腺苷（Ado）的磷酸化反应并生成一磷酸腺苷（AMP）。通过单交换介导的载体整合，在变铅青链霉菌中破坏了ADK基因（adk(Sli)，SCO2158的直系同源基因）。adk(Sli)破坏突变体（Deltaadk(Sli)）无法形成孢子，而adk(Sli)的质粒拷贝恢复了Deltaadk(Sli)的孢子形成能力，这表明adk(Sli)的缺失消除了变铅青链霉菌的孢子形成能力。添加Ado强烈抑制了变铅青链霉菌野生型（wt）的孢子形成能力，这支持了adk(Sli)的破坏导致Ado积累，进而抑制孢子形成的观点。无细胞实验表明Deltaadk(Sli)缺乏ADK活性，体外特性分析证实adk(Sli)编码ADK。adk(Sli)缺失后，Ado的细胞内水平显著升高，而AMP水平显著降低，而Deltaadk(Sli)在S-腺苷同型半胱氨酸（SAH）和S-腺苷甲硫氨酸（SAM）水平上与野生型无显著差异。值得注意的是，向野生型添加Ado会降低AMP含量，尽管未降至Deltaadk(Sli)的水平，这意味着Deltaadk(Sli)中Ado积累部分导致了AMP水平的降低。在Deltaadk(Sli)中，放线紫红素（ACT）的产生受到抑制，而十一烷基灵菌红素（RED）的产生显著增强；然而，添加Ado未能发挥这种差异控制作用。启动子探针分析证实了Deltaadk(Sli)中actII-orf4的抑制和redD的诱导，证实了ADK缺乏导致的未知代谢变化引发了变铅青链霉菌次级代谢的差异基因控制。本研究首次揭示了Ado在链霉菌发育中的抑制作用以及ADK活性在链霉菌次级代谢中的差异调节功能，尽管其潜在机制尚待阐明。
