Capone Ricardo F, Ning Yu, Pakulis Nora, Alhazzazi Turki, Fenno J Christopher
Department of Biologic and Materials Sciences, School of Dentistry, University of Michigan, Ann Arbor, MI 48109-1078, USA.
FEMS Microbiol Lett. 2007 Mar;268(2):261-7. doi: 10.1111/j.1574-6968.2006.00589.x. Epub 2006 Dec 20.
The Treponema denticola ATCC 35405 genome annotation contains most of the genes for de novo pyrimidine biosynthesis. To initiate characterization of pyrimidine synthesis in Treponema, we focused on TDE2110 (the putative pyrF, encoding orotidine-5'-monophosphate decarboxlyase). Unlike the parent strain, an isogenic pyrF mutant was resistant to 5-fluoroorotic acid. In complex medium, growth of the pyrF mutant was independent of added uracil, indicating activity of a uracil uptake/salvage pathway. Transcription of pyrF was greatly reduced in T. denticola grown in excess uracil, demonstrating that de novo pyrimidine synthesis is regulated and suggesting a feedback mechanism. Treponema denticola PyrF complemented uracil auxotrophy in an Escherichia coli pyrF mutant. This study provides biochemical confirmation of T. denticola genome predictions of de novo and salvage pyrimidine pathways and provides proof of concept that pyrF has potential as a selectable marker in T. denticola.
齿垢密螺旋体ATCC 35405的基因组注释包含了从头嘧啶生物合成的大部分基因。为了开始对齿垢密螺旋体中的嘧啶合成进行表征，我们聚焦于TDE2110（假定的pyrF，编码乳清苷-5'-单磷酸脱羧酶）。与亲本菌株不同，一个同基因的pyrF突变体对5-氟乳清酸具有抗性。在复合培养基中，pyrF突变体的生长不依赖于添加的尿嘧啶，这表明存在尿嘧啶摄取/补救途径的活性。在过量尿嘧啶中生长的齿垢密螺旋体中，pyrF的转录大幅降低，这表明从头嘧啶合成受到调控，并提示存在一种反馈机制。齿垢密螺旋体PyrF补充了大肠杆菌pyrF突变体中的尿嘧啶营养缺陷型。本研究为齿垢密螺旋体基因组中从头和补救嘧啶途径的预测提供了生化证实，并提供了pyrF在齿垢密螺旋体中具有作为选择标记潜力的概念验证。
