Biocatalytic Approaches to Building Blocks for Enzymatic and Chemical Glycan Synthesis

Abstract

While the field of biocatalysis has bloomed over the past 20−30 years, advances in the understanding and improvement of carbohydrate-active enzymes, in particular, the sugar nucleotides involved in glycan building block biosynthesis, have progressed relatively more slowly. This perspective highlights the need for further insight into substrate promiscuity and the use of biocatalysis fundamentals (rational design, directed evolution, immobilization) to expand substrate scopes toward such carbohydrate building block syntheses and/or to improve enzyme stability, kinetics, or turnover. Further, it explores the growing premise of using biocatalysis to provide simple, cost-effective access to stereochemically defined carbohydrate materials, which can undergo late-stage chemical functionalization or automated glycan synthesis/polymerization.

Realizing the Continuous Chemoenzymatic Synthesis of Anilines Using an Immobilized Nitroreductase

“…This includes transformations once thought of as impossible for enzymes, such as cyclopropanation, , C–N, C–Si, and C–C bond forming reactions . Biocatalysis has also seen an increase in application toward complex biomolecules including peptides, (oligo)­nucleotides, and carbohydrates . An area where biotransformations hold particular promise in is the direct replacement of synthetic methods that are perceived as unsustainable, in the long term .…”

The Conversion of UDP‐Glc to UDP‐Man: In Silico and Biochemical Exploration To Improve the Catalytic Efficiency of CDP‐Tyvelose C2‐Epimerases

“…Despite the good understanding of glycosyltransferases and the natural NS biosynthesis pathways, the biggest hurdle for biocatalytic in vitro glycodiversification remains the lack of (cheaply) available NSs [4,5] . Different multi‐enzymatic cascades for different common nucleotide sugars, such as UDP‐Glc, dTDP‐Glc, GDP‐Man and UDP‐GlcNAc, as well as the rare GDP‐ l ‐fucose have been described [4,5] .…”

“…Despite the good understanding of glycosyltransferases and the natural NS biosynthesis pathways, the biggest hurdle for biocatalytic in vitro glycodiversification remains the lack of (cheaply) available NSs [4,5] . Different multi‐enzymatic cascades for different common nucleotide sugars, such as UDP‐Glc, dTDP‐Glc, GDP‐Man and UDP‐GlcNAc, as well as the rare GDP‐ l ‐fucose have been described [4,5] . However, these biocatalytic pathways usually pose many challenges for large scale NS production due to many unwanted by‐products, inhibitory effects of the by‐products on the pathway enzymes, expensive downstream processing, and difficult automation lead to high process costs and overall to extremely high prices per gram of NS [4,5] …”

“…This short one‐step reaction and the application of sucrose as cheap substrate significantly lowered the production costs and made it commercially available for a few hundred dollars per gram [7] . In this context, the conversion of NDP‐Glc to NDP‐Man is very attractive as the latter is the natural starting point for the biosynthesis of many rare NSs, such as GDP‐ l ‐fucose, GDP‐perosamine and GDP‐ l ‐collitose [4,5,8] . A promiscuous CDP‐tyvelose 2‐epimerase (TyvE) from Thermodesulfatator atlanticus ( Ta TyvE) was recently found to catalyze this reaction as side activity [9] …”

“…[4,5] Different multi-enzymatic cascades for different common nucleotide sugars, such as UDP-Glc, dTDP-Glc, GDP-Man and UDP-GlcNAc, as well as the rare GDP-lfucose have been described. [4,5] However, these biocatalytic pathways usually pose many challenges for large scale NS production due to many unwanted by-products, inhibitory effects of the by-products on the pathway enzymes, expensive downstream processing, and difficult automation lead to high process costs and overall to extremely high prices per gram of NS. [4,5] UDP-Glc is the NS with the best established commercial production pipeline starting from the cheap substrate sucrose, which is converted in presence of UDP by a sucrose synthase to UDP-Glc and fructose.…”