Glycoside Hydrolase Family 98

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• Author: Fathima Shaikh
• Responsible Curator: Al Boraston

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Glycoside Hydrolase Family GH98
Clan	Not assigned
Mechanism	Inverting
Active site residues	Known
CAZy DB link
https://www.cazy.org/GH98.html

Substrate specificities

The glycoside hydrolases of this family are endo-β-galactosidases. No other activities have been reported. Family 98 glycoside hydrolases are unique in their specificity towards cleavage of the type II core [gal(β1-4)glcNAc] in the AB blood group antigens (EABase, Sp3GH98) and Lewis^Y antigens (SpGH98) [1, 2, 3]. These enzymes are capable of processing these glycans when presented on cell surfaces thus destroying the antigens [1, 2].

Kinetics and Mechanism

Family GH98 galactosidases are inverting enzymes, as first shown by NMR monitoring of the Sp4GH98-catalyzed hydrolysis of the Lewis^Y tetrasaccharide [2]. EABase was also shown to act through an inverting enzyme by NMR monitoring of the EABase catalysed hydrolysis of an artificial substrate, DNP-A-trisaccharide [3]. These results are contrary to the initial predictions made by Rigden [4]. EABase follows normal Michaelis-Menten kinetics [3].

Catalytic Residues

The general base, an aspartate and glutamate diad, and the general acid, glutamate, were identified through the crystal structures of Sp3GH98 and Sp4GH98 in complex with the A trisaccharide and H disaccharide, respectively, and confirmed by site-directed mutagenesis [2]. Similar results were obtained through kinetic studies of the corresponding acid and base mutants of EABase with the artificial substrate dinitrophenyl A-trisaccharide. Further biochemical evidence for the catalytic acid residue was obtained by comparison between the activity of the acid mutant and the pK_a of the leaving group of the substrate [3].

Three-dimensional structures

The first crystal structures from family 98 were the Sp3GH98 and Sp4GH98 enzymes from S. pneumoniae in complex with the A trisaccharide and H disaccharide respectively [2]. Both structures feature a (α/β)₈ barrel in the catalytic domain with an adjoining β-sandwich domain that contributes to the architecture of the active site and helps define the respective specificities of the enzymes [2]

Family Firsts

First sterochemistry determination

Streptococcus pneumoniae TIGR4 endo-beta-galactosidase Sp4GH98 by NMR [2].

First general base identification

Streptococcus pneumoniae SP3-BS71 endo-beta-galactosidase Sp3GH98 [2].

Streptococcus pneumoniae TIGR4 endo-beta-galactosidase Sp4GH98 [2].

First general acid identification

Streptococcus pneumoniae SP3-BS71 endo-beta-galactosidase Sp3GH98 [2].

Streptococcus pneumoniae TIGR4 endo-beta-galactosidase Sp4GH98 [2].

First 3-D structures

Streptococcus pneumoniae SP3-BS71 endo-beta-galactosidase Sp3GH98 [2].

Streptococcus pneumoniae TIGR4 endo-beta-galactosidase Sp4GH98 [2].

References

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