Role of hypoxanthine-guanine phosphoribosyltransferase in the metabolism of fairy chemicals in rice.

Fairy chemicals (FCs), 2-azahypoxanthine (AHX), imidazole-4-carboxamide (ICA), and 2-aza-8-oxohypoxanthine (AOH), are molecules with many diverse functions in plants. The defined biosynthetic pathway for FCs is a novel purine metabolism in which they are biosynthesized from 5-aminoimidazole-4-carboxamide. Here, we show that one of the purine salvage enzymes, hypoxanthine-guanine phosphoribosyltransferase (HGPRT), recognizes AHX and AOH as substrates. Two novel compounds, AOH ribonucleotide and its ribonucleoside which are the derivatives of AOH, were enzymatically synthesized. The structures were determined by mass spectrometry, 1D and 2D NMR spectroscopy, and X-ray single-crystal diffraction analysis. This report demonstrates the function of HGPRT and the existence of novel purine metabolism associated with the biosynthesis of FCs in rice.

Utilization of Corn Steep Liquor for the Production of Fairy Chemicals by Lepista sordida Mycelia.

There are various potential practical uses of fairy chemicals (FCs) in the fields of agriculture, cosmetics, and medicine; however, the production costs of FCs are very high. To enable the practical use of FCs, more efficient and inexpensive methods of culturing the mycelia of FCs-producing fungi and producing FCs need to be developed. The purpose of the present study was to determine methods of reducing the production costs of FCs and mycelia of the FCs-producing fungus Lepista sordida. We investigated the effects of four food industrial by-products, i.e., corn steep liquor (CSL), rice bran, wheat bran, and Japanese liquor lees, as nutritional additives in the liquid culture medium of the fungus. We found that CSL was more effective than the other tested additives in increasing the production of FCs and mycelia. Medium containing 1% CSL was optimal for increasing the mycelial yield while medium containing 6% CSL was optimal for increasing the production of FCs. The reason for this difference in the optimal CSL concentration was considered to be related to the stress on the mycelia caused by the amount of nutrients in the liquid medium. These results are expected to facilitate the practical use of FCs and the mycelia of FCs-producing fungi.

Biosynthesis of the Fairy Chemicals, 2-Azahypoxanthine and Imidazole-4-carboxamide, in the Fairy Ring-Forming Fungus Lepista sordida.

Fairy rings resulting from a fungus-plant interaction appear worldwide. 2-Azahypoxanthine (AHX) and imidazole-4-carboxamide (ICA) were first isolated from the culture broth of one of the fairy ring-forming fungi, Lepista sordida. Afterward, a common metabolite of AHX in plants, 2-aza-8-oxohypoxanthine (AOH), was found in AHX-treated rice. The biosynthetic pathway of the three compounds that are named as fairy chemicals (FCs) in plants has been partially elucidated; however, that in mushrooms remains unknown. In this study, it was revealed that the carbon skeletons of AHX and ICA were constructed from Gly in L. sordida mycelia and the fungus metabolized 5-aminoimidazole-4-carboxamide (AICA) to both of the compounds. These results indicated that FCs were biosynthesized by a diversion of the purine metabolic pathway in L. sordida mycelia, similar to that in plants. Furthermore, we showed that recombinant adenine phosphoribosyltransferase (APRT) catalyzed reversible interconversion not only between 5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranosyl 5'-monophosphate (AICAR) and AICA but also between ICA-ribotide (ICAR) and ICA. Furthermore, the presence of ICAR in L. sordida mycelia was proven for the first time by LC-MS/MS detection, and this study provided the first report that there was a novel metabolic pathway of ICA in which its ribotide was an intermediate in the fungus.

A Fairy Chemical, Imidazole-4-carboxamide, is Produced on a Novel Purine Metabolic Pathway in Rice.

Rings or arcs of fungus-regulated plant growth occurring on the floor of woodlands and grasslands are commonly called "fairy rings". Fairy chemicals, 2-azahypoxanthine (AHX), imidazole-4-carboxamide (ICA), and 2-aza-8-oxohypoxanthine (AOH), are plant growth regulators involved in the phenomenon. The endogeny and biosynthetic pathways of AHX and AOH in plants have already been proven, however, those of ICA have remained unclear. We developed a high-sensitivity detection method for FCs including ICA and the endogenous ICA was detected in some plants for the first time. The quantitative analysis of the endogenous level of ICA in rice and Arabidopsis were performed using 13C-double labeled ICA. In addition, the incorporation experiment and enzyme assay using the labeled compound into rice and partially purified fraction of rice indicated that ICA is biosynthesized from 5-aminoimidazole-4-carboxamide (AICA), a metabolite on the purine metabolic pathway. The relationship between ICA and AHX was also discussed based on quantitative analysis and gene expression analysis.

N-Glucosides of Fairy Chemicals, 2-Azahypoxanthine and 2-Aza-8-oxohypoxanthine, in Rice.

Plant growth stimulators, 2-azahypoxanthine (AHX) and 2-aza-8-oxohypoxanthine (AOH), were isolated from the fairy-ring-forming fungus, Lepista sordida, and AHX-treated rice, respectively. Further metabolites of AHX were detected in AHX-treated rice by HPLC, and the metabolites 1-4 were isolated from the rice. The structures of 1-4 were determined by spectroscopic analysis and synthesis. Compounds 1-4 exhibited no significant activity against rice, indicating that rice regulates the activity of AHX and AOH by converting them into their glucosides.