Manipulating the biosynthesis of glycosaminoglycans
(2025)- Abstract
- Glycosaminoglycans are structurally diverse polysaccharides that play essential roles in cellular signaling, extracellular matrix organization, and disease progression. Despite their biological importance, the mechanisms of GAG biosynthesis remain not fully understood. This thesis aims to advance mechanistic insight into the biosynthetic machinery and to develop chemical tools and inhibitors targeting two key enzymes: β4GalT7 and DS-epi1. The work was structured in two parts: probing GAG biosynthesis with functionalized xylosides, and rational design of carbohydrate-based enzyme inhibitors.
In the first part, fluorescently labeled xylosides were synthesized to investigate their potential as primers for GAG biosynthesis. While... (More) - Glycosaminoglycans are structurally diverse polysaccharides that play essential roles in cellular signaling, extracellular matrix organization, and disease progression. Despite their biological importance, the mechanisms of GAG biosynthesis remain not fully understood. This thesis aims to advance mechanistic insight into the biosynthetic machinery and to develop chemical tools and inhibitors targeting two key enzymes: β4GalT7 and DS-epi1. The work was structured in two parts: probing GAG biosynthesis with functionalized xylosides, and rational design of carbohydrate-based enzyme inhibitors.
In the first part, fluorescently labeled xylosides were synthesized to investigate their potential as primers for GAG biosynthesis. While efficiently galactosylated by β4GalT7 and internalized by cells, full-length GAG chains were not observed, highlighting limitations imposed by fluorophore charge and localization. To overcome these challenges, an azide-functionalized naphthoxyloside was developed, which primed
biosynthesis of native-like GAG chains and enabled bioorthogonal labeling through copper-free click chemistry. This probe allowed both surface immobilization and confocal imaging, providing a versatile platform for studying GAG structure and function.
The second part is focused on rational inhibitor design. A series of uridine-bisphosphonate naphthoxylosides were prepared as transition-state mimics for β4GalT7. Although the parent compounds proved unstable, the hydrolyzed product exhibited inhibitory activity (IC50 = 188 μM), suggesting a smaller, more stable scaffold for future exploration. In parallel, a library of 1,4-modified glucuronic acids was synthesized as DS-epi1 inhibitors. Screening identified a lead compound with an IC50 of 42 μM, validating the design rationale and demonstrating the enzyme’s tolerance toward anomeric modifications.
Together, these studies establish new molecular tools to probe GAG biosynthesis and provide initial scaffolds for selective enzyme inhibition. The results show both the opportunities and challenges in translating carbohydrate chemistry into functional probes and inhibitors. By expanding the chemical biology toolbox for GAG research, this work contributes to future efforts aimed at understanding and therapeutically
targeting disorders of proteoglycan metabolism. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/0de5ff30-a3ff-4d9d-8d0f-110ee943976f
- author
- Mastio, Roberto LU
- supervisor
-
- Ulf Ellervik LU
- Emil Tykesson LU
- opponent
-
- Professor Almqvist, Fredrik, Umeå Universitet
- organization
- publishing date
- 2025-11
- type
- Thesis
- publication status
- published
- subject
- keywords
- Xyloside, Glycosaminoglycan, Proteoglycan, Carbohydrate synthesis, GAG priming, GAG inhibition, β4GalT
- pages
- 84 pages
- publisher
- Lund University, Faculty of Science, Department of Chemistry, Centre for Analysis and Synthesis
- defense location
- Kemicentrum sal KC:A. Join via zoom: https://lu-se.zoom.us/j/65485564302
- defense date
- 2025-11-27 09:00:00
- ISBN
- 978-91-8096-123-3
- 978-91-8096-122-6
- language
- English
- LU publication?
- yes
- id
- 0de5ff30-a3ff-4d9d-8d0f-110ee943976f
- date added to LUP
- 2025-11-03 10:15:07
- date last changed
- 2025-11-04 10:24:34
@phdthesis{0de5ff30-a3ff-4d9d-8d0f-110ee943976f,
abstract = {{Glycosaminoglycans are structurally diverse polysaccharides that play essential roles in cellular signaling, extracellular matrix organization, and disease progression. Despite their biological importance, the mechanisms of GAG biosynthesis remain not fully understood. This thesis aims to advance mechanistic insight into the biosynthetic machinery and to develop chemical tools and inhibitors targeting two key enzymes: β4GalT7 and DS-epi1. The work was structured in two parts: probing GAG biosynthesis with functionalized xylosides, and rational design of carbohydrate-based enzyme inhibitors.<br/><br/>In the first part, fluorescently labeled xylosides were synthesized to investigate their potential as primers for GAG biosynthesis. While efficiently galactosylated by β4GalT7 and internalized by cells, full-length GAG chains were not observed, highlighting limitations imposed by fluorophore charge and localization. To overcome these challenges, an azide-functionalized naphthoxyloside was developed, which primed<br/>biosynthesis of native-like GAG chains and enabled bioorthogonal labeling through copper-free click chemistry. This probe allowed both surface immobilization and confocal imaging, providing a versatile platform for studying GAG structure and function.<br/><br/>The second part is focused on rational inhibitor design. A series of uridine-bisphosphonate naphthoxylosides were prepared as transition-state mimics for β4GalT7. Although the parent compounds proved unstable, the hydrolyzed product exhibited inhibitory activity (IC50 = 188 μM), suggesting a smaller, more stable scaffold for future exploration. In parallel, a library of 1,4-modified glucuronic acids was synthesized as DS-epi1 inhibitors. Screening identified a lead compound with an IC50 of 42 μM, validating the design rationale and demonstrating the enzyme’s tolerance toward anomeric modifications.<br/><br/>Together, these studies establish new molecular tools to probe GAG biosynthesis and provide initial scaffolds for selective enzyme inhibition. The results show both the opportunities and challenges in translating carbohydrate chemistry into functional probes and inhibitors. By expanding the chemical biology toolbox for GAG research, this work contributes to future efforts aimed at understanding and therapeutically<br/>targeting disorders of proteoglycan metabolism.}},
author = {{Mastio, Roberto}},
isbn = {{978-91-8096-123-3}},
keywords = {{Xyloside; Glycosaminoglycan; Proteoglycan; Carbohydrate synthesis; GAG priming; GAG inhibition; β4GalT}},
language = {{eng}},
publisher = {{Lund University, Faculty of Science, Department of Chemistry, Centre for Analysis and Synthesis}},
school = {{Lund University}},
title = {{Manipulating the biosynthesis of glycosaminoglycans}},
url = {{https://lup.lub.lu.se/search/files/232047463/Avhandling_Roberto_Mastio_LUCRIS.pdf}},
year = {{2025}},
}