The research outlined in this proposal is aimed at elucidating the mode of biosynthesis of a number of antibiotics. Compounds to be studied are pyrrolnitrin and the pyrrolomycins, the hydroaromatic antibiotics ketomycin, dihydrophenylalanine and anticapsin, some benzoisochromane quinone antibiotics, the macrocyclic antibiotics aplasmomycin, naphthomycin and ansatrienine, several compounds containing biosynthetically unusual structural units like asukamycin, manumycin, reductiomycin and acarbose, and the modified peptide antibiotics nosiheptide and thiostreptone. Biological structure modification based on biosynthetic information is also envisioned with some of these compounds. The methods to be used include (a) feeding experiments with precursors labeled with radioactive isotopes, but also to a large extent the use of 13C, 15N, 18O and deuterium labeled compounds in conjunction with Fourier Transform NMR Spectroscopy, (b) isolation of pathway intermediates from blocked mutants, (c) isolation of enzymes carrying out some of these transformations and (d) synthesis and use of stereospecifically labeled substrates to probe the stereochemistry and mechanism of some of the biosynthetic reactions.
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