Enzyme-Catalyzed Synthesis of Furanosyl Nucleotides

Abstract

A bacterial alpha-d-glucopyranosyl-1-phosphate thymidylyltransferase was found to couple four hexofuranosyl-1-phosphates, as well as a pentofuranosyl-1-phosphate, with deoxythymidine 5'-triphosphate, providing access to furanosyl nucleotides. The enzymatic reaction mixtures were analyzed by electrospray ionization mass spectrometry and NMR spectroscopy to determine the anomeric stereochemistry of furanosyl nucleotide products. This is the first demonstration of a nucleotidylyltransferase discriminating between diastereomeric mixtures of sugar-1-phosphates to produce stereopure, biologically relevant furanosyl nucleotides.

Stereospecific synthesis of sugar-1-phosphates and their conversion to sugar nucleotides

“…The stereospecific outcome of the phosphorylation step is well known for L L-fucose derivatives 36 where neighboring group participation controls the attack of the phosphate at the anomeric center, as only the bottom face of the compound would be open to attack in this circumstance, resulting in 1,2-trans stereochemistry. 22 Consistent with this mechanistic outcome, we present the first stereospecific preparation of a-L L-arabinose-1-phosphate (15), whereupon phosphorylation of 13 only the a anomer was prepared with an observed 3 J H-1,H-2 value of 7.5 Hz, suggesting a 4 C 1 chair conformation. The outcome of this phosphorylation method with acylated a-L L-rhamnosyl bromide (3) has previously been reported incorrectly by Zhao and Thorson, 37 where the isolation of only the b diastereomer was reported.…”

“…12 The recent observation that the glycosyltransferase VinC is capable of processing both a-and b-anomer of a sugar nucleotide to give glycosylated products with corresponding inversion of stereochemistry at the anomeric linkage 13 underlies the importance of stereospecific synthetic methods for the preparation of these substrates. Although the enzymatic preparation of these compounds has emerged, [14][15][16][17][18] the versatility available via chemical synthesis is unparalleled.…”

Hydrolysis and Formation of Glycosidic Bonds

“…pneumoniae [147,148], have been exploited successfully for the regeneration of CMPand dTDP-sugars, respectively.…”

Pathway and Enzyme Engineering and Applications for Glycodiversification

“…A series of 3-O-alkylglucose-1-phosphates were also found to be accepted as substrates, leading to the synthesis of UDP-3-O-methylglucose and UDP-3-O-dodecylglucose. The production of NDP-3-O-alkyl sugars [180] and NDP-hexofuranosyl sugars [181] by nucleotidylyltransferases further expanded the structural diversity accessible through in vitro glycodiversification.…”