Preparation of allosamidin and demethylallosamidin photoaffinity probes and analysis of allosamidin-binding proteins in asthmatic mice.

Allosamidins, metabolites of Streptomyces with strong inhibitory activities toward family 18 chitinases, show a variety of biological activities in various organisms. We prepared photoaffinity and biotinylated probes of allosamidin and demethylallosamidin, the N-demethyl derivative that shows much stronger anti-asthmatic activity than allosamidin. Mild acid hydrolysis of allosamidins afforded mono-amine derivatives, which were amidated to prepare probes with a photoactivatable aryl azide and/or biotin moieties. The derivatives with an N-acyl group at C-2 of the D-allosamine residue at the non-reducing end of allosamidins inhibited Trichoderma chitinase as strongly as the original compounds. Since the target of allosamidins in asthma is unclear, photoaffinity probes were used to analyze allosamidin-binding proteins in bronchoalveolar lavage (BAL) fluid in IL-13-induced asthmatic mice. Ym1, a chitinase-like protein, was identified as the main allosamidin-binding protein among proteins whose expression was upregulated by IL-13 in BAL fluid. Binding of allosamidins with Ym1 was confirmed by the experiments with photoaffinity probes and recombinant Ym1.