Glycoside Hydrolase Family 188

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• Author:
• Responsible Curator: Spencer Williams

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Glycoside Hydrolase Family GH188
Clan	GH-x
Mechanism	NAD-dependent hydrolysis
Active site residues	known
CAZy DB link
https://www.cazy.org/GH188.html

Substrate specificities

The glycoside hydrolases of this family are found in bacteria, algae, plants and a small number of archaea [1]. The family contains enzymes with sulfoquinovosidase activity (EC 3.2.1.199), namely the ability to cleave glycosides of 6-deoxy-6-sulfoquinovose. Sulfoquinovosidases are also found in family GH31 [2]. Enzymes of this family have the ability to cleave both α- and β-glycosides, and are dependent on an NAD⁺ cofactor.

Kinetics and Mechanism

GH188 enzymes utilize an NAD-dependent hydrolysis mechanism that proceeds through oxidation-elimination-addition-reduction steps. Exchange of the substrate C2 proton with solvent deuterium during the enzyme-catalyzed reaction was demonstrated by mass spectrometry [1]. The following chemical mechanism is proposed: (1) C3 hydride abstraction via the reduction of NAD⁺ cofactor to NADH and simultaneous oxidation of the C3 hydroxyl group; (2) α to the ketone functionality, the C2 proton is deprotonated by a general catalytic base residue; (3) cleavage of the C1-O1 bond occurs in an α,β-elimination, producing an α,β-unsaturated ketone intermediate; (4) 1,4-Michael-like addition of a water molecule at C1; and (5) reduction of the C3 carbonyl functionality by the enzyme-bound NADH generates the product.

Catalytic Residues

Catalytic residues can be inferred on the basis of X-ray crystallographic data for complexes of GH188 enzymes with NAD⁺ and sulfoquinovose [1]. Tyr136 in Arthrobacter sp. strain U41 SqgA is conserved in all family GH188 members and is located close to C2-OH, suggesting a possible role as a general base. His321 is located close to the C1-OH and may act as general acid (along with Tyr136) for the glycosidic oxygen facilitating glycosidic bond scission. The sulfonate group is recognized by a triad of amino acids: one oxygen H-bonds to Arg166 (2.6 Å), a second to Lys172 (2.9 Å), and a third to the backbone amide of Leu170 (2.8 Å).

Three-dimensional structures

Crystallographic data is available for at least two GH188 enzymes, including as complexes with NADH, and NADH plus sulfoquinovose. The 3D X-ray crystal structures include those of Flavobacterium sp. strain K172 SqgA (PDB 8QC8, 8QC2) and Arthrobacter sp. strain U41 SqgA (PDB 8QC3, 8QC6. 8QC5) [1]. Each enzyme possesses an N-terminal dinucleotide-binding Rossman fold. The GH188 enzymes show structural similarities to inositol-2-dehydrogenase, glucose-fructose/IDH/MocA-like oxidoreductase, and NAD⁺-dependent N-acetylgalactosaminidase of family GH109.

Family Firsts

First stereochemistry determination

not applicable

First catalytic residue identification

Arthrobacter sp. strain U41 SqgA using 3D X-ray crystal structure [1]

First 3-D structural determination

Flavobacterium sp. strain K172 SqgA and Arthrobacter sp. strain U41 SqgA [1]

First GH188 enzyme shown to hydrolyze both α- and β-substrates

Flavobacterium sp. strain K172 SqgA [1]

1. Kaur2024 pmid=38100472

1. Speciale2016 pmid=26878550

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