Insights on DHAP Aldolases ability to convert Dioxygen or a Ketone as Electrophile: use of a Strain Depleted in Triose Phosphate Isomerase

Abstract

AbstractRhamnulose‐1‐phosphate aldolases (RhuA) were the first dihydroxyacetone phosphate (DHAP) aldolases to be demonstrated to efficiently accept a ketone as the electrophile, providing the highly sought‐after chiral tertiary alcohol motif. It has also been found that this enzyme family can use dioxygen as an electrophile, forcing to work under inert atmosphere conditions when the target electrophile is poorly reactive. However, these features have not been explored in all DHAP‐aldolase families. Here, we present a study of E. coli K12 fuculose‐1‐phosphate‐ (FucAK12), fructose‐1,6‐bisphosphate‐ (FruAK12) and tagatose‐1,6‐bisphosphate (TagAK12) aldolases. For the latter two enzymes, this was made possible by the development of a triose phosphate isomerase (TPI)‐deleted E coli strain, which prevents DHAP isomerization into d‐glyceraldehyde 3‐phosphate (G3P), its natural electrophile. Like RhuA, TagAK12 was found to be able to oxidize DHAP into hydroxypyruvaldehyde phosphate under dioxygen, leading to the corresponding diulose. On the other hand, FucAK12 was revealed to be capable of converting ketone electrophiles with greater stereoselectivity than RhuA, and without the requirement of an inert atmosphere, since it was unable to oxidize DHAP. Finally, FruAK12 proved unable either to oxidize DHAP and or to accept ketones as electrophiles.