Engineering a Fungal L-Arabinose Pathway Towards the Co-Utilization of Hemicellulosic Sugars for Production of Xylitol
type: CreativeWork
description: The biosynthesis of value-added products has shown great promise in recent years due to the advances in molecular biology and protein engineering. Many advantages over chemical synthesis include high selectivity and specificity, increased yield, and function under milder conditions without the need for toxic metals, organic solvents, strong acids or bases, or high pressures and temperatures. However, naturally occurring enzymatic pathways are often times not particularly well suited towards obtaining high product yield due to the organism's evolution not dependent on the production of such high levels of desired product. Xylitol is an attractive pentose sugar alcohol that has many applications in the food, pharmaceutical, and high-value based biochemical product industries. It is still, however, relatively expensive to produce, which prevents its economical integration into the marketplace. To expand the possible routes of xylitol synthesis, and provide a process which can utilize both hemicellulosic pentose sugars D-xylose and L-arabinose from renewable plant biomass as feed substrates, my thesis research focused on utilizing a fungal-derived biosynthetic pathway for production of xylitol. The pathway consists of two NADPH-dependent reductases, xylose reductase (XR) and L-xylulose reductase (LXR), and an NAD+-dependent L-arabinitol 4-dehydrogenase (LAD). The XR enzyme converts D-xylose directly to xylitol, while the three enzyme in tandem convert L-arabinose to xylitol. However, the cofactor imbalance presented with the pathway potentially makes nicotinamide regeneration difficult for production in vivo. After cloning and characterization of a highly active and stable NAD +-dependent L-arabinitol 4-dehydrogenase (LAD) from Neurospora crassa, subsequent engineering via rational design and directed evolution resulted in the isolation of the first known NADP+-dependent LAD enzyme. This novel engineered LAD was then introduced into the fungal xylitol pathway and expres…
abstract: The biosynthesis of value-added products has shown great promise in recent years due to the advances in molecular biology and protein engineering. Many advantages over chemical synthesis include high selectivity and specificity, increased yield, and function under milder conditions without the need for toxic metals, organic solvents, strong acids or bases, or high pressures and temperatures. However, naturally occurring enzymatic pathways are often times not particularly well suited towards obtaining high product yield due to the organism's evolution not dependent on the production of such high levels of desired product. Xylitol is an attractive pentose sugar alcohol that has many applications in the food, pharmaceutical, and high-value based biochemical product industries. It is still, however, relatively expensive to produce, which prevents its economical integration into the marketplace. To expand the possible routes of xylitol synthesis, and provide a process which can utilize both hemicellulosic pentose sugars D-xylose and L-arabinose from renewable plant biomass as feed substrates, my thesis research focused on utilizing a fungal-derived biosynthetic pathway for production of xylitol. The pathway consists of two NADPH-dependent reductases, xylose reductase (XR) and L-xylulose reductase (LXR), and an NAD+-dependent L-arabinitol 4-dehydrogenase (LAD). The XR enzyme converts D-xylose directly to xylitol, while the three enzyme in tandem convert L-arabinose to xylitol. However, the cofactor imbalance presented with the pathway potentially makes nicotinamide regeneration difficult for production in vivo. After cloning and characterization of a highly active and stable NAD +-dependent L-arabinitol 4-dehydrogenase (LAD) from Neurospora crassa, subsequent engineering via rational design and directed evolution resulted in the isolation of the first known NADP+-dependent LAD enzyme. This novel engineered LAD was then introduced into the fungal xylitol pathway and expres…
author: Sullivan, Ryan Patrick
keywords: Engineering, Chemical
date published: 2014-12-17T20:39:26Z
date modified: 2026-07-22T22:26:06Z
provider: University of Illinois - Urbana-Champaign
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Illinois at Urbana-Champaign

PropertyValue
property ID: Handle
value: http://hdl.handle.net/2142/72139