Abstract
Allosamidin 1 is a first chitinase inhibitor produced by Streptomyces sp. It possesses a novel pseudotrisaccharide structure which consists of N-acetyl-D-allosamine and allosamizoline 2. This has a novel cyclopentanoid structure which is highly oxygenated and fused with a dimethylaminooxazoline ring. In this study, we elucidated the biosynthesis of 1. First, the origin of the carbon skeleton and nitrogen atoms was elucidated by means of feeding experiments with labelled precursors. Incorporation experiments using [1-^<13>C]- and [6-^<13>C]glucose as well as doubly labelled [1-^<13>C,2-^<15>N]glucosamine revealed that each skeleton of N-acetylallosamine and 2 was derived from D-glucosamine. Further experiments with L-[guanidino-^<13>C,^<15>N]arginine and [methyl-^<13>C]methionine clarified the origin of the dimethylaminooxazoline moiety of 2. Next, ^<14>C-labelled 1, 3, 4, 5, 6 and 7 were prepared to investigate the biosynthesis of 1. Conversion experiments with the labelled allosamidins revealed that 3 was a precursor of 1, but 4 was not incorporated. This suggests that the first N-methyl group is introduced before the cyclization of the aminooxazoline ring during the biosynthesis of 1. Although none of the compounds 5, 6 and 7 were converted to 1, the production of 1 was inhibited by the addition of 5. To study a mechanism of the cyclopentane ring formation of 1, feeding experiments with [4-^2H], [5-^2H] and [6-^2H_2]-D-glucosamine were carried out. ^2H NMR analysis of the labeled samples indicated that loss of deuterium on C-5 and stereospecific loss of one of the two deuterium on C-6 of glucosamine occured during the biosynthesis of the cyclopentane ring of 1. These results suggested that the cyclization to form the cyclopentanoid moiety of 1 would proceed via an intermediate 6-aldehyde glucosamine derivative.
