Direct Evidence That 8-Dehydrogenation Channels the Biosynthetic Pathway of Rifamycins

Area of Science:

• Biochemistry
• Molecular Biology
• Natural Product Biosynthesis

Background:

• The 8-hydroxy (8-OH) group of rifamycin is crucial for its biological activity.
• The mechanism of naphthalene ring formation involving the 8-OH group in rifamycin biosynthesis was previously unclear.

Purpose of the Study:

• To elucidate the biochemical and structural basis of 8-dehydrogenation in rifamycin biosynthesis.
• To propose a pathway for the conversion of prorifamycin A to 34a-deoxyrifamycin W.

Main Methods:

• Biochemical assays to analyze enzyme activity.
• Structural analysis of enzyme complexes.
• Proposed biosynthetic pathway reconstruction.

Main Results:

• Identified a rare S2T2 complex of NAD+-dependent dehydrogenases, RifS and RifT.
• RifS catalyzes 8-dehydrogenation via a canonical NAD+-dependent mechanism.
• RifT functions as a noncatalytic partner in the complex.
• Proposed a pathway from benzene-type prorifamycin A to naphthalene-type 34a-deoxyrifamycin W.

Conclusions:

• Provided direct evidence for the branch point in rifamycin and 8-deoxyrifamycin biosynthesis.
• Established the role of RifS and RifT in 8-dehydrogenation.
• Opened avenues for engineering novel ansamycin compounds.