The Flavoenzyme Azobenzene Reductase AzoR from Escherichia coli Binds Roseoflavin Mononucleotide (RoFMN) with High Affinity and Is Less Active in Its RoFMN Form

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• 作者：Simone Langer ; Shinobu Nakanishi ; Tilo Mathes ; Tanja Knaus ; Alexandra Binter ; Peter Macheroux ; Tomoko Mase ; Takuya Miyakawa ; Masaru Tanokura ; Matthias Mack
• 刊名：Biochemistry
• 出版年：2013
• 出版时间：June 25, 2013
• 年：2013
• 卷：52
• 期：25
• 页码：4288-4295
• 全文大小：390K
• 年卷期：v.52,no.25(June 25, 2013)
• ISSN：1520-4995

文摘

The Gram-positive bacterium Streptomyces davawensis is the only organism known to produce the antibiotic roseoflavin. Roseoflavin is a structural riboflavin analogue and is converted to the flavin mononucleotide (FMN) analogue roseoflavin mononucleotide (RoFMN) by flavokinase. FMN-dependent homodimeric azobenzene reductase (AzoR) (EC 1.7.1.6) from Escherichia coli was analyzed as a model enzyme. In vivo and in vitro experiments revealed that RoFMN binds to the AzoR apoenzyme with an even higher affinity compared to that of the 鈥渘atural鈥?cofactor FMN. Structural analysis (at a resolution of 1.07 脜) revealed that RoFMN binding did not affect the overall topology of the enzyme and also did not interfere with dimerization of AzoR. The AzoR鈥揜oFMN holoenzyme complex was found to be less active (30% of AzoR鈥揊MN activity) in a standard assay. We provide evidence that the different physicochemical properties of RoFMN are responsible for its reduced cofactor activity.