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Related Experiment Videos

Direct evidence that the rifamycin polyketide synthase assembles polyketide chains processively.

T W Yu1, Y Shen, Y Doi-Katayama

  • 1Department of Chemistry, Box 351700, University of Washington, Seattle, WA 98195-1700, USA.

Proceedings of the National Academy of Sciences of the United States of America
|August 4, 1999
PubMed
Summary

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The rifF gene product terminates polyketide synthase assembly, releasing rifamycin B. Disruptions reveal linear polyketides, indicating processive PKS action and early oxidative cyclization during assembly.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Microbiology

Background:

  • Rifamycin B is a crucial antibiotic produced by Amycolatopsis rifamycinica.
  • Polyketide synthases (PKS) are large multi-enzyme complexes responsible for synthesizing polyketide natural products.
  • The biosynthesis of rifamycin B involves a type I PKS encoded by rifA-rifE genes.

Purpose of the Study:

  • To investigate the role of the rifF gene in rifamycin B biosynthesis.
  • To understand the processivity and termination mechanisms of the rifamycin polyketide synthase.
  • To elucidate the timing and nature of oxidative cyclization during polyketide assembly.

Main Methods:

  • Gene inactivation studies (mutant generation).
  • Analysis of accumulated polyketide intermediates using structural elucidation techniques.

Related Experiment Videos

  • Isotope labeling experiments ((18)O) to track oxygen incorporation.

    • Main Results:

    • Inactivation of rifF leads to the accumulation of linear polyketides, confirming PKS processivity.
    • Premature termination of polyketide assembly by disrupting rifD, rifE module 9, or module 10 also results in linear polyketide accumulation.
    • Oxidative cyclization to a naphthoquinone structure occurs during, not after, PKS assembly, forming an 8-hydroxy-7,8-dihydronaphthoquinone intermediate.

    Conclusions:

    • The rifF gene product is essential for the final macrocyclization and release of rifamycin B.
    • The rifamycin PKS operates in a processive manner, assembling extended polyketide chains before termination.
    • Oxidative modifications, including cyclization to naphthoquinone structures, are integrated into the PKS assembly pathway.