In conclusion, we characterized the equilibrium between ribosyl nucleosides and their corresponding 2′,3′-borate esters in aqueous solution, a phenomenon which facilitates apparent equilibrium shifts during nucleoside phosphorolysis and glycosylation reactions due to a biased esterification of nucleosides over the sugar phosphate Rib1P. This borate esterification also causes decreases of the phosphorolysis rate by pyrimidine nucleoside phosphorylases, most likely via non-productive binding of the nucleoside borate ester to the enzyme and its hydrolytic interconversion to the free substrate. Collectively, the effects described herein shine light on the activity of nucleoside-binding enzymes in the presence of borate and provide an orthogonal dimension for reaction engineering in nucleobase glycosylation reactions.