Natural low-molecular mass organic compounds with oxidase activity as organocatalysts

Abstract

Organocatalysts, low-molecular mass organic compounds composed of nonmetallic elements, are often used in organic synthesis, but there have been no reports of organocatalysts of biological origin that function in vivo. Here, we report that actinorhodin (ACT), a natural product derived from Streptomyces coelicolor A3(2), acts as a biocatalyst. We purified ACT and assayed its catalytic activity in the oxidation of L-ascorbic acid and L-cysteine as substrates by analytical methods for enzymes. Our findings were as follows: (i) oxidation reactions producing H 2 O 2 proceeded upon addition of ACT to the reaction mixture; (ii) ACT was not consumed during the reactions; and (iii) a small amount (catalytic amount) of ACT consumed an excess amount of the substrates. Even at room temperature, atmospheric pressure, and neutral pH, ACT showed catalytic activity in aqueous solution, and ACT exhibited substrate specificity in the oxidation reactions. These findings reveal ACT to be an organocatalyst. ACT is known to show antibiotic activity, but its mechanism of action remains unknown. On the basis of our results, we propose that ACT kills bacteria by catalyzing the production of toxic levels of H 2 O 2 . We also screened various other natural products of bacterial, plant, and animal origins and found that several of the compounds exhibited catalytic activity, suggesting that living organisms produce and use these compounds as biocatalysts in nature.A catalyst is a substance that increases the rate of a chemical reaction but is not consumed in the reaction. Catalysts are classified as homogeneous or heterogeneous, and most of the major homogeneous catalysts are metal-containing catalysts. In general, enzymes and ribozymes, which are classified as homogeneous catalysts, are referred to as biocatalysts. Enzymes are large proteins that catalyze numerous reactions in living organisms whereas ribozymes are composed of RNA and are known to catalyze phosphoryl transfer reactions (involved in RNA selfsplicing), the formation of peptide bonds, and various other reactions (1-5). Organocatalysts, which are homogeneous catalysts, are low-molecular mass organic compounds derived from nonmetallic elements (e.g., carbon, oxygen, hydrogen, and nitrogen) (6). Organocatalysts are used in industry because they have the following advantages over metal-containing catalysts (7-10): (i) Organocatalysts exhibit catalytic activity under mild conditions (atmospheric pressure, room temperature, and neutral pH); (ii) disposal of depleted organocatalysts is inexpensive; (iii) environmental loads due to waste reaction mixtures containing organocatalysts are low; and (iv) the risk of product contamination by metal ions is low. Therefore, organocatalysts have been receiving much attention in the field of green chemistry (11).In our laboratory, we have been studying the metabolism of actinomycetes from both fundamental and applied points of view (12)(13)(14)(15). From Streptomyces, we discovered a previously unidentified enzyme, L-glutamate oxidase (EC 1.4.3...

Actinorhodin is a redox‐active antibiotic with a complex mode of action against Gram‐positive cells

“…In agreement with previous work, we found that actinorhodin had antibiotic activity against Staphylococcus aureus , although it is sixfold more potent than previously reported with an MIC of 4 µg ml −1 rather than 25–30 µg ml −1 (Wright and Hopwood, ). Consistent with its spectral responses to pH (Bystrykh et al ., ; Nishiyama et al ., ), we found that its antibacterial activity was also pH‐responsive. The compound was most potent under acidic conditions, with a MIC of 0.25 µg ml −1 at pH 5.5 (Fig.…”

“…Alternatively, the discrepancy could be due to the difference in methodology where the previous report extrapolated actinorhodin's MIC using an agar plug assay. The fact that actinorhodin is more potent at low pH argues against the biological relevance of its in vitro organocatalyst activity, as this effect is more pronounced in alkaline pH (Nishiyama et al ., ).…”

“…coelicolor (Dietrich et al ., ; Dela Cruz et al ., ; Shin et al ., ; Singh et al ., ; Lee et al ., ). Third, actinorhodin has been shown to act as an organocatalyst of oxidative reactions in vitro , catalyzing the conversion of l ‐ascorbic acid and l ‐cysteine with the concomitant production of H 2 O 2 (Nishiyama et al ., ). In that work, it was suggested that actinorhodin might kill bacteria via the accumulation of toxic concentrations of H 2 O 2 .…”

Discovery of a sesamin-metabolizing microorganism and a new enzyme

“…Unique pathways of microbial metabolism of these compounds have been reported (30)(31)(32)(33)(34). Research on the microbial metabolism of a diversity of natural compounds can be expected to reveal novel enzymes (4) and catalytic functions (34).…”

Total Synthesis of Actinorhodin

“…As the 1 H NMR spectrum of the natural product had not been reported, we conducted the fermentation of Streptomyces coelicolor A3(2) in our laboratory and isolated an authentic sample of 1 according to the reported procedure . Upon direct comparison, the 1 H NMR spectra of the synthetic and authentic specimens fully coincided.…”