Publications

• Microbial degradation of validamycin A by Flavobacterium saccharophilum. Enzymatic cleavage of C-N linkage in validoxylamine A.

  Asano N., Takeuchi M., Ninomiya K., Kameda Y., Matsui K.

  The enzymatic cleavage of C-N linkage in the degradation of validamycin A by Flavobacterium saccharophilum was examined using N-p-nitrophenyl derivatives of validamine and valienamine as synthetic model substrates for validoxylamine A. Incubation of N-p-nitrophenylvalidamine with the membrane frac ... >> More

  The enzymatic cleavage of C-N linkage in the degradation of validamycin A by Flavobacterium saccharophilum was examined using N-p-nitrophenyl derivatives of validamine and valienamine as synthetic model substrates for validoxylamine A. Incubation of N-p-nitrophenylvalidamine with the membrane fraction from the organism led to formation of N-p-nitrophenyl-3-ketovalidamine, and succeeding cleavage of C-N linkage. As the products of the cleavage step, one was identified as p-nitroaniline and another keto compound could not be purified enough because of its instability. However, on the basis of its hydrogenation products, the structure of the keto compound could be established as 5D-(5/6)-5-C-(hydroxy-methyl)-2,6-dihydroxy-2-cyclohexen-1-one. The same experiment was carried out with N-p-nitrophenylvalienamine. In this case, N-p-nitrophenyl-3-ketovalienamine could be isolated as an intermediate but the desired keto compound from the cleavage step could not be isolated because of its instability. The participation of two enzymes, that is, a dehydrogenase and a C-N lyase on the cleavage of C-N linkage was assured, and moreover, the analysis of its products, together with those of the previous studies allow us to propose a degradation pathway of validamycin A by Flavobacterium saccharophilum. << Less

  J Antibiot (Tokyo) 37:859-867(1984) [PubMed] [EuropePMC]

• Purification and properties of 3-ketovalidoxylamine A C-N lyase from Flavobacterium saccharophilum.

  Takeuchi M., Asano N., Kameda Y., Matsui K.

  3-Ketovalidoxylamine A C-N lyase was purified about 900-fold from the cell-free extract of Flavobacterium saccharophilum by ammonium sulfate fractionation, column chromatography on CM cellulose and gel filtration on Sephacryl S-200. The purified enzyme was homogeneous as judged by sodium dodecyl s ... >> More

  3-Ketovalidoxylamine A C-N lyase was purified about 900-fold from the cell-free extract of Flavobacterium saccharophilum by ammonium sulfate fractionation, column chromatography on CM cellulose and gel filtration on Sephacryl S-200. The purified enzyme was homogeneous as judged by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis. The molecular weight of the enzyme was estimated to be 36,000 by gel filtration on Sephacryl S-200 and by SDS polyacrylamide gel electrophoresis, indicating that the enzyme is a monomer. The optimum pH was found at 9.0. The enzyme activity was inhibited by EDTA or ethyleneglycol bis(beta-aminoethylether)-N,N'-tetraacetic acid and the inhibition was reversed by Ca2+ ion. The enzyme was able to eliminate p-nitroaniline or p-nitrophenol from p-nitrophenyl-3-ketovalidamine (IV) or p-nitrophenyl-alpha-D-3-ketoglucoside (VI), but not from p-nitrophenyl-1-epi-3-ketovalidamine or p-nitrophenyl-beta-D-3-ketoglucoside. Apparent Km values for IV and VI were 0.24 mM and 0.5 mM, respectively. << Less

  J Biochem 98:1631-1638(1985) [PubMed] [EuropePMC]