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The novel phosphatase RosC catalyzes the last unknown step of roseoflavin biosynthesis in Streptomyces davaonensis.
Molecular Microbiology ( IF 3.6 ) Pub Date : 2020-07-04 , DOI: 10.1111/mmi.14567
Carmen Schneider 1 , Valentino Konjik 1 , Lena Kißling 1 , Matthias Mack 1
Affiliation  

The bacterium Streptomyces davaonensis produces the antibiotic roseoflavin, which is a riboflavin (vitamin B2) analog. The key enzyme of roseoflavin biosynthesis is the 8‐demethyl‐8‐amino‐riboflavin‐5ʹ‐phosphate (AFP) synthase RosB which synthesizes AFP from riboflavin‐5ʹ‐phosphate. AFP is not a substrate for the last enzyme of roseoflavin biosynthesis the N, N‐dimethyltransferase RosA, which generates roseoflavin from 8‐demethyl‐8‐amino‐riboflavin (AF). Consequently, the roseoflavin biosynthetic pathway depends on a phosphatase, which dephosphorylates AFP to AF. Here, we report on the identification and characterization of such an AFP phosphatase which we named RosC. The gene rosC is located immediately downstream of rosA and both genes are part of a cluster comprising 10 genes. Deletion of rosC from the chromosome of S. davaonensis led to reduced roseoflavin levels in the corresponding recombinant strain. In contrast to wild‐type S. davaonensis, cell‐free extracts of the rosC deletion strain did not catalyze dephosphorylation of AFP. RosC was purified from an overproducing Escherichia coli strain. RosC is the fastest enzyme of roseoflavin biosynthesis (kcat 31.3 ± 1.4 min–1). The apparent KM for the substrate AFP was 34.5 µM. Roseoflavin biosynthesis is now completely understood––it takes three enzymes (RosB, RosC, and RosA) to convert the flavin cofactor riboflavin‐5ʹ‐phosphate into a potent antibiotic.

中文翻译:

新型磷酸酶RosC催化达瓦链霉菌中玫瑰黄素生物合成的最后一个未知步骤。

细菌链霉菌davaonensis产生抗生素玫瑰黄色素,这是一种核黄素(维生素B 2)的模拟。玫瑰黄素生物合成的关键酶是8-去甲基-8-氨基核黄素-5ʹ-磷酸(AFP)合酶RosB,它由核黄素5ʹ-磷酸合成AFP。AFP不是玫瑰黄素生物合成的最后一个酶NN-二甲基转移酶RosA的底物,后者从8-去甲基-8-氨基核黄素(AF)生成玫瑰黄素。因此,玫瑰黄素的生物合成途径取决于磷酸酶,磷酸酶将AFP磷酸化为AF。在这里,我们报告了这种命名为RosC的AFP磷酸酶的鉴定和表征。rosC基因该基因位于rosA的下游,并且两个基因都是包含10个基因的簇的一部分。从达沃链霉菌的染色体中删除rosC导致相应重组菌株中玫瑰黄素水平降低。与野生型达沃链球菌相比rosC缺失菌株的无细胞提取物不催化AFP的去磷酸化。从过量生产的大肠杆菌菌株中纯化RosC 。RosC是玫瑰黄素生物合成最快的酶(k cat 31.3±1.4 min –1)。表观K M基板AFP的λ为34.5 µM。玫瑰黄素的生物合成现已被完全理解-需要三种酶(RosB,RosC和RosA)将黄素辅因子核黄素5lav-磷酸转化为有效的抗生素。
更新日期:2020-07-04
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