Glycoside Hydrolase Family 78

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Author: ^^^Zui Fujimoto^^^
Responsible Curator: ^^^Zui Fujimoto^^^

Glycoside Hydrolase Family GH78
Clan	GH-M
Mechanism	inverting
Active site residues	known
CAZy DB link
http://www.cazy.org/GH78.html

Substrate specificities

Genes encoding family GH78 glycoside hydrolases are found in bacteria and fungi. The sole identified activity of enzymes of this family is hydrolysis of α-L-rhamnosides (EC 3.2.1.40). The GH78 α-L-rhamnosidases catalyze the hydrolysis of α-L-rhamnosyl-linkages in L-rhamnosides, including: flavonoid glycosides such as naringin, hesperidin and rutin; polysaccharides such as rhamnogalacturonan and arabinogalactan-protein and glycolipids. α-L-Rhamnosidases have been found to be one component of rhamnogalacturonan hydrolase [1], or naringinase [2].

Kinetics and Mechanism

GH78 enzymes hydrolyze glycosidic bonds through an inverting mechanism as elucidated by proton NMR [3, 4]. Typical GH78 α-L-rhamnosidases have molecular masses in the range 80-120 kDa, and are most active at pH 4.0 to 8 and temperature of 50°C against p-nitrophenyl-α-L-rhamnopyranoside [1, 5, 6, 7, 8].

Catalytic Residues

Crystallographic and mutagenesis studies of Streptomyces avermitilis α-L-rhamnosidase (SaRha78A), notably including an enzyme-product complex structure, suggested that Glu895 is the catalytic general base responsible for activating a water molecule, and that Glu636 is the catalytic general acid, assisting leaving-group departure [9]. All characterized α-L-rhamnosidases appear to contain a glutamate as the catalytic general base.

Three-dimensional structures

The first crystal structure of a GH78 member was determined for Bacillus sp. GL1 α-L-rhamnosidase B (BsRhaB) (PDB ID 2okx) [10]. Subsequently, the crystal structure of the putative α-L-rhamnosidase BT1001 from Bacteroides thetaiotaomicron VPI-5482 was determined by a structural genomics project (PDB ID 3cih) [11]. The crystal structure of Streptomyces avermitilis α-L-rhamnosidase (SaRha78A) in complex with the product L-rhamnose has revealed key active-site details (PDB IDs 3w5m, 3w5n) [9]. More recently, the crystal structure of a Klebsiella oxytoca α-rhamnosidase (KoRha) has been solved in complex L-rhamnose.

α-L-Rhamnosidases have a modular structure. BsRhaB, BT1001, and SaRha78A show five-, four and six-module structures. The catalytic domain of GH78 enzymes is an (α/α)₆-barrel. A fibronectin type-3 fold β-domain often appears at the N-terminus, and a C-terminal Greek key β-domain exists just after the catalytic domain. Several β-domains are also inserted between the N-terminal domain and the catalytic domain. Streptomyces avermitilis α-L-rhamnosidase (SaRha78A) possesses one carbohydrate binding module (CBM67), which binds terminal L-rhamnose sugars in the presence of a calcium ion [9]. On the other hand, KoRha has only two structual domains, one β-domain and one catalytic domain, forming a homodimer [12]. These two domains are common among all structure-determined enzymes.

Family Firsts

First stereochemistry determination: Aspergillus aculeatus α-L-rhamnosidase (RhaA), by ¹H-NMR [3].
First general base residue identification: Streptomyces avermitilis α-L-rhamnosidase (SaRha78A), based on mutagensis informed by 3D structural data [9].
First general acid residue identification: Streptomyces avermitilis α-L-rhamnosidase (SaRha78A), based on mutagensis informed by 3D structural data [9].
First 3-D structure: Bacillus sp. GL1 α-L-rhamnosidase B (BsRhaB) (PDB IDs 2okx) [10].

References

All Medline abstracts: PubMed