Changes in biotin levels during production of natto, Japanese fermented soybean

<p><span style="font-family: Times New Roman;"><span style="font-size: medium;">The change of biotin level during production of natto (Japanese fermented soybean) was investigated in this study. </span><span style="font-size: medium;">The total biotin level was measured by an agar plate bioassay using </span><em><span style="font-size: medium;">Lactobacillus plantarum</span></em><span style="font-size: medium;"> ATCC 8014.</span><span style="font-size: medium;"> </span><span style="font-size: medium;">The total biotin level decreased during water soaking, but increased after the fermentation of soybeans using </span><em><span style="font-size: medium;">Bacillus subtilis</span></em><span style="font-size: medium;"> var. </span><em><span style="font-size: medium;">natto</span></em><span style="font-size: medium;"> (</span><em><span style="font-size: medium;">B. natto</span></em><span style="font-size: medium;">) and reached a maximum level.</span><span style="font-size: medium;"> </span><span style="font-size: medium;">The increase of total biotin was not affected by Asp, Arg, and Ile which promoted the growth of </span><em><span style="font-size: medium;">L. plantarum</span></em><span style="font-size: medium;"> in high concentrations. </span><span style="font-size: medium;"> </span><span style="font-size: medium;">The peak level of biotin in the fermented soybeans was significantly higher than that of dry soybeans.</span><span style="font-size: medium;"> </span><span style="font-size: medium;">The fermented soybeans at the biotin peak level were adequate for food.</span><span style="font-size: medium;"> </span><span style="font-size: medium;">In addition, we detected 9 and 4 biotinylated polypeptides in the soybeans and </span><em><span style="font-size: medium;">B. natto</span></em><span style="font-size: medium;"> used in this study, respectively.</span><span style="font-size: medium;"> </span><span style="font-size: medium;">We speculated that the increase of biotin level may depend on the increase of the 4 biotinylated polypeptides and free biotin in </span><em><span style="font-size: medium;">B. natto</span></em><span style="font-size: medium;">.</span></span></p>

An Extracellular Protease of Pseudomonas fluorescens Inactivates Antibiotics of Pantoea agglomerans

Pseudomonas fluorescens A506 and Pantoea agglomerans strains Eh252 and C9-1 are biological control agents that suppress fire blight, an important disease of pear and apple caused by the bacterium Erwinia amylovora. Pseudomonas fluorescens strain A506 suppresses disease largely through competitive exclusion of E. amylovora on surfaces of blossoms, the primary infection court, whereas Pantoea agglomerans strains Eh252 and C9-1 produce antibiotics that are toxic to E. amylovora. In this study, an extracellular protease produced by A506 is characterized and evaluated for its capacity to inactivate the antibiotics produced by the strains of Pantoea agglomerans. Activity of the extracellular protease was optimal at pH 9 and inhibited by zinc- or calcium-chelators, indicating that the protease is an alkaline metalloprotease. In an agar plate bioassay, partially purified extracellular protease inactivated the antibiotics mccEh252 and herbicolin O, which are produced by Pantoea agglomerans strains Eh252 and C9-1, respectively. Derivatives of A506 deficient in extracellular protease production were obtained by transposon mutagenesis, and the aprX gene encoding the protease was cloned and sequenced. Strain A506 inactivated mccEh252 and herbicolin O in agar plate bioassays, whereas the aprX mutant did not inactivate the antibiotics. Both A506 and the aprX mutant were insensitive to antibiosis by C9-1 and Eh252; thus, the protease was not required to protect A506 from antibiosis. These data highlight a previously unknown role of the extracellular protease produced by Pseudomonas fluorescens A506 in interactions among plant-associated microbes.