Functional characterization and engineering of the sugar donor specificity of a 2″-O-xylosyltransferase from Panax notoginseng for rare saponins biosynthesis

  • Int J Biol Macromol. 2026 May:362:152095. doi: 10.1016/j.ijbiomac.2026.152095.
Maoqi Hou  1 ,  Xunjiang Wang  1 ,  Sijie Chen  1 ,  Wenxiang Fan  1 ,  Li Yang  2 ,  Shujuan Zhao  3 ,  Zhengtao Wang  4
Affiliations
  • 1. State Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, The SATCM Key Laboratory for New Resources & Quality Evaluation of Chinese Medicine, The MOE Key Laboratory for Standardization of Chinese Medicines and Shanghai Key Laboratory of Compound Chinese Medicines, Institute of Chinese Materia Medica, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China.
  • 2. State Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, The SATCM Key Laboratory for New Resources & Quality Evaluation of Chinese Medicine, The MOE Key Laboratory for Standardization of Chinese Medicines and Shanghai Key Laboratory of Compound Chinese Medicines, Institute of Chinese Materia Medica, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China. Electronic address: [email protected].
  • 3. State Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, The SATCM Key Laboratory for New Resources & Quality Evaluation of Chinese Medicine, The MOE Key Laboratory for Standardization of Chinese Medicines and Shanghai Key Laboratory of Compound Chinese Medicines, Institute of Chinese Materia Medica, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China. Electronic address: [email protected].
  • 4. State Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, The SATCM Key Laboratory for New Resources & Quality Evaluation of Chinese Medicine, The MOE Key Laboratory for Standardization of Chinese Medicines and Shanghai Key Laboratory of Compound Chinese Medicines, Institute of Chinese Materia Medica, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China. Electronic address: [email protected].
PMID: 42002181 DOI: 10.1016/j.ijbiomac.2026.152095
Abstract

2″-O-xylosylation is a rare glycosylation pattern conferring unique bioactivities to triterpenoid saponins in Panax species. Here, we identify a novel 2″-O-xylosyltransferase, PnUGT94X1, from P. notoginseng, which regioselectively catalyzes 2″-O-xylosylation at the C3-linked sugar of diverse saponins. PnUGT94X1 exhibits a pronounced preference for UDP-xylose and a broad specificity tolerance for sugar acceptors including protopanaxadiol-, cycloartenol-, and oleanane-type saponins. Site-directed mutagenesis revealed that the residues C140 and T141, with a distance of 22 residues downstream of the Asp in the catalytic His-Asp dyad, synergistically govern substrate preference between UDP-xylose and UDP-glucose, which is defined as an important part of sugar donor selectivity-determining region (SDSR). Using PnUGT94X1 and its engineered mutants, eleven rare saponins including six new compounds were biosynthesized by 2″-O-xylosylation or 2″-O-glucosylation, three of them exhibited significant effects (at 5 mg·kg-1) against LPS-induced acute lung injury in mice. The elucidated SDSR module provides a strategic framework for engineering UGTs to alter their sugar donors, thereby enabling the synthesis structurally diverse glycosides.

Keywords
Engineering; Glycosyltransferase; Panax notoginseng; Saponin; Sugar donor.
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