7F7R

Enterococcus faecalis GH31 alpha-N-acetylgalactosaminidase D455N in complex with Tn antigen

Summary for 7F7R

• Entry DOI: 10.2210/pdb7f7r/pdb
• Descriptor: GH31 alpha-N-acetylgalactosaminidase, 2-acetamido-2-deoxy-alpha-D-galactopyranose, SERINE, ... (7 entities in total)
• Functional Keywords: glycoside hydrolase, gh31, mucin, (beta/alpha)8-barrel, fibronectin-like, substrate, complex, hydrolase
• Biological source: Enterococcus faecalis ATCC 10100
• Total number of polymer chains: 1
• Total formula weight: 107889.02

Authors

Miyazaki, T. (deposition date: 2021-06-30, release date: 2021-12-08, Last modification date: 2023-11-29)

Primary citation

Miyazaki, T.,Ikegaya, M.,Alonso-Gil, S.
Structural and mechanistic insights into the substrate specificity and hydrolysis of GH31 alpha-N-acetylgalactosaminidase.
Biochimie, 195:90-99, 2022

PubMed Abstract: Glycoside hydrolase family 31 (GH31) is a diversified family of anomer-retaining α-glycoside hydrolases, such as α-glucosidase and α-xylosidase, among others. Recently, GH31 α-N-acetylgalactosaminidases (Nag31s) have been identified to hydrolyze the core of mucin-type O-glycans and the crystal structure of a gut bacterium Enterococcus faecalis Nag31 has been reported. However, the mechanisms of substrate specificity and hydrolysis of Nag31s are not well investigated. Herein, we show that E. faecalis Nag31 has the ability to release N-acetylgalactosamine (GalNAc) from O-glycoproteins, such as fetuin and mucin, but has low activity against Tn antigen. Mutational analysis and crystal structures of the Michaelis complexes reveal that residues of the active site work in concert with their conformational changes to act on only α-N-acetylgalactosaminides. Docking simulations using GalNAc-attached peptides suggest that the enzyme mainly recognizes GalNAc and side chains of Ser/Thr, but not strictly other peptide residues. Moreover, quantum mechanics calculations indicate that the enzyme preferred p-nitrophenyl α-N-acetylgalactosaminide to Tn antigen and that the hydrolysis progresses through a conformational itinerary, C → S → C, in GalNAc of substrates. Our results provide novel insights into the diversification of the sugar recognition and hydrolytic mechanisms of GH31 enzymes.

PubMed: 34826537
DOI: 10.1016/j.biochi.2021.11.007

Experimental method

X-RAY DIFFRACTION (1.63 Å)