8XC8

beta-1,4-galacosyltransferase

Summary for 8XC8

• Entry DOI: 10.2210/pdb8xc8/pdb
• Descriptor: Glycosyltransferase family 25 protein (2 entities in total)
• Functional Keywords: transferase
• Biological source: Aggregatibacter actinomycetemcomitans NUM4039
• Total number of polymer chains: 2
• Total formula weight: 59684.05

Authors

Luo, G.,Huang, Z.,Chen, J.,Hou, X.,Zhu, Y.,Ni, D.,Xu, W.,Zhang, W.,Rao, Y.,Mu, W. (deposition date: 2023-12-08, release date: 2024-12-11)

Primary citation

Luo, G.,Huang, Z.,Zhu, Y.,Chen, J.,Hou, X.,Ni, D.,Xu, W.,Zhang, W.,Rao, Y.,Mu, W.
Crystal structure and structure-guided tunnel engineering in a bacterial beta-1,4-galactosyltransferase.
Int.J.Biol.Macromol., 279:135374-135374, 2024

PubMed Abstract: Lacto-N-neotetraose (LNnT), a representative oligosaccharide found in human milk, has been previously examined for its beneficial traits. However, the LNnT titer is limited by the efficient glycosyltransferase pathway, particularly with respect to the catalysis of rate-limiting steps. As data on the crystal structure of the key enzyme required for synthesizing LNnT are lacking, the synthesis of LNnT remains an uncertainty. Here, for the first time we report the three-dimensional structure of a bacterial β-1,4-galactosyltransferase, Aaβ4GalT, and analyze the critical role played by residues in its catalytic efficacy. Guided by structural insights, we engineered this enzyme to enhance its catalytic efficiency using structure-guided tunnel engineering. The mutant enzyme L5 (K155M/H156D/F157W/K185M/Q216V) so produced, showed a 50-fold enhancement in catalytic activity. Crystal structure analysis revealed that the mechanism underlying the improvement in activity was of the swing door type. The closed conformation formed by dense hydrophobic packing with Q216V-K155M widened and permitted substrate entry. Our results show that altering the tunnel conformation helped appropriately accommodate the substrate for catalysis and provide a structural basis for the modification of other glycosyltransferases.

PubMed: 39265897
DOI: 10.1016/j.ijbiomac.2024.135374

Experimental method

X-RAY DIFFRACTION (2.51 Å)