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Glycoside Hydrolase Family 123
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- Author: ^^^Tomomi Sumida^^^
- Responsible Curator: ^^^Tomomi Sumida^^^
| Glycoside Hydrolase Family GH123 | |
| Clan | GH-x |
| Mechanism | retaining/inverting |
| Active site residues | known/not known |
| CAZy DB link | |
| https://www.cazy.org/GH123.html | |
Substrate specificities
The glycoside hydrolases family 123 contains β-N-acetylgalactosaminidases (EC 3.2.1.53). These enzymes specifically hydrolyze the non-reducing terminal β-GalNAc linkage, but not β-GlcNAc linkage. The β-N-acetylgalactosaminidase (EC 3.2.1.53) is distinguished from β-hexosaminidase (EC 3.2.1.52) or β-N-acetylglucosaminidase (EC 3.2.1.52). Because the β-N-acetylgalactosaminidase is specific to β-GalNAc linkage while β-N-acetylglucosaminidase is specific to β-GlcNAc linkage. β-Hexosaminidase hydrolyzes both β-GlcNAc and β-GalNAc linkages at non-reducing terminal. NgaP, N-acetylgalactosaminidase from Paenibacillus sp., is the first cloned β-N-acetylgalactosaminidase and its primary structure is not similar to any glycohydrolases reported so far [1], and, thus, this family is created. The recombinant NgaP hydrolyzes pNP-β-GalNAc but not pNP-β-GlcNAc, pNP-β-Gal, pNP-α-GalNAc or other pNP-glycosides, indicating that NgaP is a typical β-N-acetylgalactosaminidase.
Kinetics and Mechanism
Glycoside hydrolases belonging to GH18, GH20 and GH85 cleave the sugar containing C2-acetamide group such as β-GlcNAc and β-GalNAc through substrate-assisted catalysis involving neighboring group participation. Since NgaP hydrolyzes the β-GalNAc linkage, NgaP is proposed to use substrate-assisted catalysis. A comparison of secondary structure of NgaP with that of other enzymes that utilize substrate-assisted catalysis suggested that Glu608 of NgaP functions as a proton donor. Point mutation analysis confirmed that Glu608 is integral for the activity of NgaP. GalNAc-thiazoline, a structural analog of the oxazolinium intermediate of neighboring group participation, was found to competitively inhibit the activity of NgaP. These results indicate that NgaP hydrolyzes the terminal β-GalNAc linkage through substrate-assisted catalysis.
Catalytic Residues
Point mutation analysis suggested that Glu608 functions as a proton donor in NgaP.
Three-dimensional structures
Unknown
Family Firsts
- First stereochemistry determination
- First catalytic nucleophile identification
The carbonyl oxygen of the C-2 acetamide group of the substrate behaves as a catalytic nucleophile.
- First general acid/base residue identification
Site-directed mutagenesis indicated that Glu608 is an essential amino acid for the catalytic reaction in NgaP.
- First 3-D structure
Not known