Glycoside Hydrolase Family 112

• Author: Motomitsu Kitaoka
• Responsible Curator: Shinya Fushinobu

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Glycoside Hydrolase Family 112
Clan	none
Mechanism	inverting
Active site residues	known
CAZy DB link
http://www.cazy.org/fam/GH112.html

Substrate specificities

This family contains beta-galactoside phosphorolyzing enzymes, β-1,3-D-galactosyl-D-hexososamine phosphorylase [1] and β-1,4-D-gaactosyl-L-rhamnose phosphorylase [2]. The former enzymes are subcategorized into galacto-N-biose phosphorylase, (GNBP) [3], lacto-N-biose I phosphorylase (LNBP) [4], and galacto-N-biose/lacto-N-biose I phosphorylase (GLNBP) [1, 5, 6] based on the substrate preference on galacto-N-biose (GNB, Gal-β1,3-GalNAc) and lacto-N-biose I (LNB, Gal-β1,3-GlcNAc) [4].

Kinetics and Mechanism

Phosphorolysis by GH112 enzymes proceeds with inversion of anomeric configuration, as first shown by Derensy-Dron et al. [1] on β-1,3-D-galactosyl-D-hexososamine phosphorylase from Bifidobacterium bifidum, i.e. LNB + Pi ↔ α-galactose 1-phosphate + GlcNAc. Considering the topology of the active site structure, the reaction mechanism for inverting phosphorylase is proposed to be similar to that for inverting GH. With the aid of general acid residue, the enzymatic phosphorolysis begins with direct nucleophilic attack by phosphate on the anomeric C-1 carbon, instead of the water molecule activated by a general base residue in inverting GH reaction.

Catalytic Residues

The catalytic proton donar residue was firstly estimated by mutational analysis on β-1,3-D-galactosyl-D-hexososamine phosphorylase from Bifidobacterium longum [6]. Catalytic nucleophile is phosphate.

Three-dimensional structures

The first solved 3-D structure was β-1,3-D-galactosyl-D-hexososamine phosphorylase from Bifidobacterium longum [7]. The catalytic domain consists of a partially broken TIM barrel fold that is structurally similar to a β-galactosidase of GH42, supporting the classification of these phosphorolytic enzymes as one of the GH families. A large conformational change in TIM barrel scaffold was observerd with substrate-binding.

Family Firsts

First sterochemistry determination

β-1,3-D-galactosyl-D-hexososamine phosphorylase from Bifidobacterium bifidum [1].

First cloned

β-1,3-D-galactosyl-D-hexososamine phosphorylase from Bifidobacterium longum [7].

First catalytic nucleophile identification

phosphate

First general acid/base residue identification

β-1,3-D-galactosyl-D-hexososamine phosphorylase from Bifidobacterium longum [8].

First 3-D structure

β-1,3-D-galactosyl-D-hexososamine phosphorylase from Bifidobacterium longum [9].

References

1. Hidaka M, Nishimoto M, Kitaoka M, Wakagi T, Shoun H, and Fushinobu S. (2009). The crystal structure of galacto-N-biose/lacto-N-biose I phosphorylase: a large deformation of a TIM barrel scaffold. J Biol Chem. 2009;284(11):7273-83. DOI:10.1074/jbc.M808525200 |