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Assaying for Inorganic Polyphosphate in Bacteria
Published on: January 21, 2019
In vitro phosphinate methylation by PhpK from Kitasatospora phosalacinea.
Williard J Werner1, Kylie D Allen, Kaifeng Hu
1School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Pullman, Washington 99164, United States.
Biochemistry
|September 29, 2011
Summary
Researchers purified and demonstrated the activity of PhpK, a P-methyltransferase enzyme. This enzyme catalyzes a key methylation step in phosalacine biosynthesis, forming a unique carbon-phosphorus-carbon bond.
Area of Science:
- Biochemistry
- Enzymology
- Natural Product Biosynthesis
Background:
- Radical S-adenosyl-L-methionine (SAM) and cobalamin-dependent methyltransferases are proposed to methylate unreactive atoms in antibiotic biosynthesis.
- No such enzymes have been previously purified or shown to be active in vitro.
Purpose of the Study:
- To purify and characterize the P-methyltransferase enzyme, PhpK, from Kitasatospora phosalacinea.
- To demonstrate the in vitro activity of PhpK in the biosynthesis of phosalacine.
Main Methods:
- Purification of the PhpK enzyme.
- In vitro enzymatic assays using methylcobalamin and a phosphinothricin precursor.
Main Results:
- PhpK was successfully purified and shown to be active in vitro.
- PhpK catalyzes the methylation of 2-acetylamino-4-hydroxyphosphinylbutanoate (N-acetyldemethylphosphinothricin) using methylcobalamin.
- The product of the reaction is 2-acetylamino-4-hydroxymethylphosphinylbutanoate (N-acetylphosphinothricin).
- This reaction creates the only known natural carbon-phosphorus-carbon linkage.
Conclusions:
- PhpK is the first purified and characterized cobalamin-dependent methyltransferase involved in antibiotic biosynthesis.
- The enzyme plays a crucial role in forming the unique C-P-C bond in phosalacine.
- This study provides significant insights into the mechanisms of organophosphorus compound biosynthesis.
