Direct Evidence That the Rifamycin Polyketide Synthase Assembles Polyketide Chains Processively

The assembly of the polyketide backbone of rifamycin B on the type I rifamycin polyketide synthase (PKS), encoded by the rifA-rifE genes, is terminated by the product of the rifF gene, an amide synthase that releases the completed undecaketide as its macrocyclic lactam. Inactivation of rifF gives a...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1999-08, Vol.96 (16), p.9051-9056
Main Authors: Yu, Tin-Wein, Shen, Yuemao, Doi-Katayama, Yukiko, Tang, Li, Park, Cheonseok, Moore, Bradley S., Hutchinson, C. Richard, Floss, Heinz G.
Format: Article
Language:English
Subjects:
Actinomycetales - enzymology
Actinomycetales - genetics
Amino Acid Sequence
Base Sequence
Biochemistry
Biological Sciences
Biosynthesis
Chromophores
DNA
Enzymes
Genes
Molecular Sequence Data
Molecular Structure
Multienzyme Complexes - genetics
Multienzyme Complexes - metabolism
Mutagenesis
Naphthoquinones
Polyketides
Polymerase chain reaction
Restriction Mapping
rifamycin polyketide synthase
Rifamycins
Rifamycins - biosynthesis
Rifamycins - metabolism
rifD gene
rifE gene
rifF gene
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Summary:The assembly of the polyketide backbone of rifamycin B on the type I rifamycin polyketide synthase (PKS), encoded by the rifA-rifE genes, is terminated by the product of the rifF gene, an amide synthase that releases the completed undecaketide as its macrocyclic lactam. Inactivation of rifF gives a rifamycin B nonproducing mutant that still accumulates a series of linear polyketides ranging from the tetra- to a decaketide, also detected in the wild type, demonstrating that the PKS operates in a processive manner. Disruptions of the rifD module 8 and rifE module 9 and module 10 genes also result in accumulation of such linear polyketides as a consequence of premature termination of polyketide assembly. Whereas the tetraketide carries an unmodified aromatic chromophore, the penta- through decaketides have undergone oxidative cyclization to the naphthoquinone, suggesting that this modification occurs during, not after, PKS assembly. The structure of one of the accumulated compounds together with 18O experiments suggests that this oxidative cyclization produces an 8-hydroxy-7,8-dihydronaphthoquinone structure that, after the stage of proansamycin X, is dehydrogenated to an 8-hydroxynaphthoquinone.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.96.16.9051