A bifunctional Pasteurella multocida β1-3-galactosyl/ N-acetylgalactosaminyltransferase (PmNatB) for the highly efficient chemoenzymatic synthesis of disaccharides

Abstract

Glycosyltransferases are nature's key biocatalysts for the formation of glycosidic bonds. Discovery and characterization of new synthetically useful glycosyltransferases are critical for the development of efficient enzymatic and chemoenzymatic strategies for producing complex carbohydrates and glycoconjugates. Herein we report the identification of Pasteurella multocida PmNatB as a bifunctional single-catalytic-domain glycosyltransferase with both β1-3-galactosyltransferase and β1-3-N-acetylgalactosaminyltransferase activities. It is a novel glycosyltransferase for constructing structurally diverse GalNAcβ3Galα/βOR and Galβ3GalNAcα/βOR disaccharides in one-pot multienzyme systems with in situ generation of UDP-sugars.

Figure 1.

Overlay of the AlphaFold-predicted structure of PmNatB (golden) and the β4GalNAcT domain of the chondroitin polymerase EcK4CP crystal structure (PDB ID: 2Z87, cyan). D95, D97, and D194 (labeled black) in PmNatB aligned well to D239, D241, and D362 (labelled red) in EcK4CP, respectively. Mn²⁺ is shown in purple. The carbons and the bonds in UDP-GalNAc are shown in pink.

Scheme 1.

Application of His₆-PmNatB in one-pot multienzyme (OPME) synthesis of Galβ3GalNAcα/βProNHCbz (3a/b) and GalNAcβ3Galα/βProNHCbz (4a/b) with in-situ generation of UDP-Gal and UDP-GalNAc, respectively.