ネクトリシンの生合成機構の解明と製法構築への応用に関する研究

広島大学(Hiroshima University)博士(農学)Doctor of Agriculturedoctora

Diversity of Nitrite Reductase Genes in “Candidatus Accumulibacter phosphatis”-Dominated Cultures Enriched by Flow-Cytometric Sorting▿

“Candidatus Accumulibacter phosphatis” is considered a polyphosphate-accumulating organism (PAO) though it has not been isolated yet. To reveal the denitrification ability of this organism, we first concentrated this organism by flow cytometric sorting following fluorescence in situ hybridization (FISH) using specific probes for this organism. The purity of the target cells was about 97% of total cell count in the sorted sample. The PCR amplification of the nitrite reductase genes (nirK and nirS) from unsorted and sorted cells was performed. Although nirK and nirS were amplified from unsorted cells, only nirS was detected from sorted cells, indicating that “Ca. Accumulibacter phosphatis” has nirS. Furthermore, nirS fragments were cloned from unsorted (Ba clone library) and sorted (Bd clone library) cells and classified by restriction fragment length polymorphism analysis. The most dominant clone in clone library Ba, which represented 62% of the total number of clones, was not found in clone library Bd. In contrast, the most dominant clone in clone library Bd, which represented 59% of the total number of clones, represented only 2% of the total number of clones in clone library Ba, indicating that this clone could be that of “Ca. Accumulibacter phosphatis.” The sequence of this nirS clone exhibited less than 90% similarity to the sequences of known denitrifying bacteria in the database. The recovery of the nirS genes makes it likely that “Ca. Accumulibacter phosphatis” behaves as a denitrifying PAO capable of utilizing nitrite instead of oxygen as an electron acceptor for phosphorus uptake

MOESM1 of Characterization of a novel oxidase from Thelonectria discophora SANK 18292 involved in nectrisine biosynthesis

Additional file 1: Fig. S1. Conversion of 4-amino-4-deoxyarabinitol to nectrisine with His-tagged NecC expressed by recombinant E. coli. a, b, and c, Absorbance chromatograms; d, e, and f, the extracted MS chromatograms for m/z 297.1 which is the [M+H]+ ion for reduced and NBD-labeled nectrisine. Conversions of 4-amino-4-arabinitol in the presence (a and d) or absence (b and e) of the recombinant NeccC are shown. For reference, chromatograms of reduced and NBD-labeled nectrisine which is equal to NBD-labeled 1,4-Dideoxy-1,4-imino-D-arabinitol are depicted in c and f