Synthesis of biolabile thioalkyl-protected phosphates from an easily accessible phosphotriester precursor

Abstract

Phosphates are regularly incorporated into bioactive small molecules, for example in sugar-1-phosphate derivatives that are used for metabolic oligosaccharide engineering. To enable efficient cellular uptake, the phosphate groups are commonly masked with biolabile S-acyl-2-thioethyl (SATE) protecting groups that are removed once the molecule is inside the cell. Typically, SATE-protected monophosphates are synthesised through phosphoramidite chemistry, which suffers from issues with hazardous and unstable reagents and can give unreliable yields. Here, we report the development of an alternative approach that makes use of an easy to synthesise tri(2-bromoethyl)phosphotriester precursor, providing access to bis-SATE-protected mixed phosphotriesters in two steps. We demonstrate the viability of our strategy on tetrabenzylated glucose as a model monosaccharide, onto which a bis-SATE-protected phosphate is introduced at either the anomeric position or at C6. We also show compability with various protecting groups and further explore the scope and limitations of the approach on different substrates, including N-acetylhexosamines and amino acid derivatives.

Xylosyltransferase Bump-and-hole Engineering to Chemically Manipulate Proteoglycans in Mammalian Cells

“…48 While we and others have made sugar-1-phosphates caged as labile bis-S-acetylthioethyl (SATE) phosphotriesters, synthesis of SATE-caged 6AzGlc-1-phosphate failed in our hands. 42,49,50 Instead, we took inspiration from the increasingly popular protide technology that has gained attention to cage phosphates in antiviral nucleotides. 51,52 To our knowledge, such chemistry has not been applied to sugar-1-phosphates yet, but for instance to sugar-6-phosphates.…”

“…49 While we and others have made sugar-1-phosphates caged as labile bis-S-acetylthioethyl (SATE) phosphotriesters, synthesis of SATE-caged 6AzGlc-1-phosphate failed in our hands. 43,50,51 Instead, we took inspiration from the increasingly popular protide technology that has gained attention to cage phosphates in antiviral nucleotides, 52,53 and has been recently used to cage sugar-phosphates. [54][55][56] We synthesized phosphoramidate diester 1 as a caged sugar-1phosphate to be deprotected by hydrolases in the cytosol of living mammalian cells (Fig.…”