Structural analysis of actinorhodin polyketide ketoreductase: cofactor binding and substrate specificity

Area of Science:

• Biochemistry
• Structural Biology
• Natural Products Chemistry

Background:

• Aromatic polyketides are crucial natural products with pharmaceutical applications.
• Polyketide synthases (PKS) are responsible for their biosynthesis.
• Ketoreductases (KR) introduce regio- and stereochemical diversity through polyketide chain reduction.

Purpose of the Study:

• To elucidate the structure and function of actinorhodin polyketide ketoreductase (act KR).
• To understand the molecular basis of polyketide biosynthesis specificity.
• To provide insights for designing novel aromatic polyketide natural products.

Main Methods:

• X-ray crystallography to solve cocrystal structures of act KR with NADP(+) and NADPH.
• Structural comparisons with fatty acid KRs.
• Active site analysis and identification of a proton-relay network.
• Acyl carrier protein and substrate docking models.

Main Results:

• Determined high-resolution structures of act KR bound to NADP(+) and NADPH, revealing a conserved Rossmann fold.
• Identified subtle structural differences between act KR and fatty acid KRs influencing substrate binding.
• Proposed a catalytic mechanism involving water molecules, NADPH, and an active site tetrad.
• Docking models illuminated the basis for KR regio- and stereoselectivity.

Conclusions:

• The determined structures provide a foundation for understanding aromatic PKS.
• Structural flexibility in the alpha6-alpha7 loop region was observed.
• These findings enhance the potential for designing novel aromatic polyketide natural products with tailored reduction patterns.