Structural Basis of Polyketide Synthase O-Methylation.

Area of Science:

• Biochemistry
• Molecular Biology
• Natural Product Synthesis

Background:

• Modular type I polyketide synthases (PKSs) are crucial for synthesizing complex natural products.
• PKS O-methyltransferases (O-MTs) are hypothesized to methylate hydroxyl or keto groups but lack structural and functional data.
• Understanding PKS O-MTs is key to unlocking novel polyketide biosynthesis pathways.

Purpose of the Study:

• To determine the domain boundaries, catalytic activity, and structure of StiD and StiE O-MTs.
• To investigate the substrate stereospecificity of PKS O-MTs.
• To provide a novel chemoenzymatic tool for polyketide modification.

Main Methods:

• X-ray crystallography to determine O-MT structures.
• Site-directed mutagenesis to identify key catalytic residues.
• Biochemical assays to characterize enzyme activity and substrate specificity.

Main Results:

• The domain boundaries and crystal structures of StiD and StiE O-MTs were determined.
• StiD O-MT demonstrated substrate stereospecificity in methylating β-hydroxyl groups.
• Key catalytic residues were identified and validated through mutagenesis and cross-validation with CurL O-MT.

Conclusions:

• The study provides the first structural and biochemical characterization of PKS O-MTs.
• These enzymes represent a new class of chemoenzymatic tools for selective modification of hydroxyl groups in polyketide biosynthesis.
• This work opens avenues for engineering novel polyketide structures.