Discovery and characterization of chondroitinase A with specificity for 4-O-sulfated GalNAc in chondroitin sulfate/dermatan sulfate
- 1National Glycoengineering Research Center, State Key Laboratory of Microbial Technology, Shandong Center of Technology Innovation for Carbohydrate, Shandong Key Laboratory of Carbohydrate and Carbohydrate-conjugate Drugs, Shandong University, Qingdao, People's Republic of China
Abstract
Chondroitin sulfate (CS) and dermatan sulfate (DS) are glycosaminoglycans whose diverse biological functions are largely determined by specific sulfation patterns. The GlcA/IdoA-GalNAc4S (A/iA unit), where GlcA, IdoA, GalNAc and 4S denote D-glucuronic acid, L-iduronic acid, GalNAc, and 4-O-sulfation, respectively, is the most prevalent sulfation motif in animal tissues and participates in a broad spectrum of physiological and pathological processes. However, chondroitinase A (CSase A), a lyase capable of selectively degrading CS/DS rich in A/iA units, has not yet been identified, which has limited our understanding of the functional roles of such structures in CS/DS. Here, we report the first discovery of a CSase A from the metagenome of sediments near the Haima cold seep. This enzyme belongs to polysaccharide lyase family 8 and exhibits strict selectivity for A/iA units, yielding disaccharides composed almost exclusively of the 4-O-sulfated disaccharide, with no other sulfated disaccharide variants detectable even under exhaustive digestion. Structural analysis of resistant tetrasaccharides and hexasaccharides, and degradation assays with structure-defined oligosaccharides demonstrate that CSase A exclusively cleaves the β-1,4 glycosidic bond preceding a disaccharide unit containing GalNAc or GalNAc4S, thereby defining its rigorous substrate specificity. Site-directed mutagenesis and activity profiling identified Glu-579, Arg-480, Tyr-427, and His-420 as residues essential for catalysis, supporting a proposed tyrosine-histidine-mediated acid-base mechanism. The discovery of CSase A provides a unique molecular tool for dissecting the functional roles of A/iA units in CS/DS-related biology and disease.
Related
- Characterization of dermatan sulfate and chondroitin sulfate profiles during brain regeneration in Styela plicata
- Structural characterization the LlaI anti-phage defense system reveals insights into the evolution of nucleotide specificity and the organization of DNA binding in McrBC restriction complexes
- Short-term cortical stimulation combined with targeted rehabilitation and Chondroitinase ABC improves upper limb function following a cervical contusion injury
- Experimental characterization of zeta viruses and discovery of betaviroid-like RNAs expand the diversity of coding circular RNA replicons
- Heparan Sulfate, Chondroitin Sulfate, and Keratan Sulfate as Regulators of Tumor Malignancy
BibTeX
@article{wang2026discovery,
title = {{Discovery and characterization of chondroitinase A with specificity for 4-O-sulfated GalNAc in chondroitin sulfate/dermatan sulfate}},
author = {Wenshuang Wang and Haixin Yi and Ping’en Zi and Xu Wang and Rouyi Zou and Yu-zhong Zhang and Fuchuan Li},
journal = {The Journal of Biological Chemistry},
year = {2026},
month = sep,
doi = {10.1016/j.jbc.2026.113594},
url = {https://doi.org/10.1016/j.jbc.2026.113594},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Discovery and characterization of chondroitinase A with specificity for 4-O-sulfated GalNAc in chondroitin sulfate/dermatan sulfate
AU - Wenshuang Wang
AU - Haixin Yi
AU - Ping’en Zi
AU - Xu Wang
AU - Rouyi Zou
AU - Yu-zhong Zhang
AU - Fuchuan Li
PY - 2026
DA - 2026/09/22
JO - The Journal of Biological Chemistry
DO - 10.1016/j.jbc.2026.113594
UR - https://doi.org/10.1016/j.jbc.2026.113594
AB - Chondroitin sulfate (CS) and dermatan sulfate (DS) are glycosaminoglycans whose diverse biological functions are largely determined by specific sulfation patterns. The GlcA/IdoA-GalNAc4S (A/iA unit), where GlcA, IdoA, GalNAc and 4S denote D-glucuronic acid, L-iduronic acid, GalNAc, and 4-O-sulfation, respectively, is the most prevalent sulfation motif in animal tissues and participates in a broad spectrum of physiological and pathological processes. However, chondroitinase A (CSase A), a lyase capable of selectively degrading CS/DS rich in A/iA units, has not yet been identified, which has limited our understanding of the functional roles of such structures in CS/DS. Here, we report the first discovery of a CSase A from the metagenome of sediments near the Haima cold seep. This enzyme belongs to polysaccharide lyase family 8 and exhibits strict selectivity for A/iA units, yielding disaccharides composed almost exclusively of the 4-O-sulfated disaccharide, with no other sulfated disaccharide variants detectable even under exhaustive digestion. Structural analysis of resistant tetrasaccharides and hexasaccharides, and degradation assays with structure-defined oligosaccharides demonstrate that CSase A exclusively cleaves the β-1,4 glycosidic bond preceding a disaccharide unit containing GalNAc or GalNAc4S, thereby defining its rigorous substrate specificity. Site-directed mutagenesis and activity profiling identified Glu-579, Arg-480, Tyr-427, and His-420 as residues essential for catalysis, supporting a proposed tyrosine-histidine-mediated acid-base mechanism. The discovery of CSase A provides a unique molecular tool for dissecting the functional roles of A/iA units in CS/DS-related biology and disease.
SN - 0021-9258
SN - 1083-351X
ER -