Investigation of carbonyl amidation and O-methylation during biosynthesis of the pharmacophore pyridyl of antitumor piericidins

Abstract

Piericidins are a large family of bacterial α-pyridone antibiotics with antitumor activities such as their anti-renal carcinoma activity exhibited recently in nude mice. The backbones of piericidins are derived from β, δ-diketo carboxylic acids, which are offloaded from a modular polyketide synthase (PKS) and putatively undergo a carbonyl amidation before α-pyridone ring formation. The tailoring modifications to the α-pyridone structure mainly include the verified hydroxylation and O-methylation of the C-4′ position and an unidentified C-5′ O-methylation. Here, we describe a piericidin producer, terrestrial Streptomyces conglobatus, which contains a piericidin biosynthetic gene cluster in two different loci. Deletion of the amidotransferase gene pieD resulted in the accumulation of two fatty acids that should be degraded from the nascent carboxylic acid released by the PKS, supporting the carbonyl amidation function of PieD during α-pyridone ring formation. Deletion of the O-methyltransferase gene pieB1 led to the production of three piericidin analogues lacking C-5′ O-methylation, therefore confirming that PieB1 specifically catalyses the tailoring modification. Moreover, bioactivity analysis of the mutant-derived products provided clues regarding the structure-function relationship for antitumor activity. The work addresses two previously unidentified steps involved in pyridyl pharmacophore formation during piericidin biosynthesis, facilitating the rational bioengineering of the biosynthetic pathway towards valuable antitumor agents.

Keywords

• Polyketide
• Pyridine
• Polyketide synthase (PKS)
• Natural product biosynthesis