(S)-α-Amino-β-hydroxyketones are important building blocks for sphingosine and sphingolipid analogs. Non-natural sphingolipid analogs have been characterized as critical biological agents with anti-cancer activity. Thus, research on the synthesis of enantiomerically pure α-amino-β-hydroxyketones can benefit pharmaceutical research. The synthesis of these ketones via a one-step biocatalytic process could replace conventional methods, which require multiple reactions and purification steps. Serine palmitoyltransferase (SPT) catalyzes the condensation reaction between L-serine and long-chain acyl-CoAs to produce (S)-α-amino-β-hydroxyketones. The irreversibility of the enzyme reaction makes SPT an excellent candidate for the direct synthesis of (S)-α-amino-β-hydroxyketones. However, SPT shows a preference for long-chain acyl-CoAs. To expand the acyl-CoA substrate scope of this enzyme, we performed site-saturation mutagenesis on an arginine which is thought to act as a gate on the substrate channel. One of the mutants showed higher activity and a greater preference toward acyl-CoAs with short alkyl chains when compared to wild-type SpSPT.

Expanding the substrate scope of serine palmitoyltransferase utilizing mutagenesis and high-throughput screening

2019 Spring National Meeting

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