Glycoside Hydrolase Family 78

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

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

Substrate specificities

Family GH78 glycoside hydrolases are found in bacteria and fungi. The characterized activity of this family is α-L-rhamnosidase (EC 3.2.1.40). α-L-Rhamnosidases catalyze the hydrolysis of α-L-rhamnosyl-linkages in L-rhamnose containing compounds, flavonoid glycosides such as naringin, hesperidin and rutin, polysaccharides such as rhamnogalacturonan and arabinogalactan-protein, or glycolipids.

α-L-Rhamnosidases have been found to be one components of rhamnogalacturonan hydrolase [1], or naringinase [].

Kinetics and Mechanism

GH78 enzymes hydrolyze glycosidic bonds through an acid base-assisted single displacement or inverting mechanism elucidated by proton NMR [2, 3].

α-L-rhamnosidases have molecular masses of 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, 4, 5, 6, 7].

Catalytic Residues

The crystallographic and mutagenesis studies of Streptomyces avermitilis α-L-rhamnosidase (SaRha78A) indicated that Glu895 appeared to be the catalytic general base, and Glu636 appeared to comprise the catalytic proton donor (acid) of the enzyme, activating a water molecule [8]. Glutamate is conserved for the catalytic general base in all characterized α-L-rhamnosidases.

Three-dimensional structures

The first crystal structure was determined for Bacillus sp. GL1 α-L-rhamnosidase B (BsRhaB) [9]. Then, crystal structure of the putative α-L-rhamnosidase BT1001 from Bacteroides thetaiotaomicron VPI-5482 was determined by Structural genom project [10]. Recently, crystal structure of Streptomyces avermitilis α-L-rhamnosidase (SaRha78A) in complex with L-rhamnose was reported [8].

α-L-Rhamnosidases have a modular structure. BsRhaB, BT1001, and SaRha78A show five-, four and six-module structures. The catalytic module of GH78 enzymes is an (α/α)₆-barrel. A fibronectin type 3 fold β-domain often appears in the N-terminus, and the Greek key β-domain exist just after the catalytic module comprising the C-terminus. Several β-domains are inserted between the N-terminal domain and the catalytic module. Streptomyces avermitilis α-L-rhamnosidase (SaRha78A) possesses one carbohydrate binding module (CBM67), which binds terminal L-rhamnose sugars in the presence of calcium ion [8].

Family Firsts

First stereochemistry determination

Aspergillus aculeatus α-L-rhamnosidase (RhaA), by ¹H-NMR [2].

First general base residue identification

Streptomyces avermitilis α-L-rhamnosidase (SaRha78A), based on mutagensis informed by 3D structural data [8].

First general acid residue identification

Streptomyces avermitilis α-L-rhamnosidase (SaRha78A), based on mutagensis informed by 3D structural data [8].

First 3-D structure

Bacillus sp. GL1 α-L-rhamnosidase B (BsRhaB) [9].

References

1. Mutter M, Beldman G, Schols HA, and Voragen AG. (1994). Rhamnogalacturonan alpha-L-rhamnopyranohydrolase. A novel enzyme specific for the terminal nonreducing rhamnosyl unit in rhamnogalacturonan regions of pectin. Plant Physiol. 1994;106(1):241-50. DOI:10.1104/pp.106.1.241 |
2. Pitson SM, Mutter M, van den Broek LA, Voragen AG, and Beldman G. (1998). Stereochemical course of hydrolysis catalysed by alpha-L-rhamnosyl and alpha-D-galacturonosyl hydrolases from Aspergillus aculeatus. Biochem Biophys Res Commun. 1998;242(3):552-9. DOI:10.1006/bbrc.1997.8009 |
3. Zverlov VV, Hertel C, Bronnenmeier K, Hroch A, Kellermann J, and Schwarz WH. (2000). The thermostable alpha-L-rhamnosidase RamA of Clostridium stercorarium: biochemical characterization and primary structure of a bacterial alpha-L-rhamnoside hydrolase, a new type of inverting glycoside hydrolase. Mol Microbiol. 2000;35(1):173-9. DOI:10.1046/j.1365-2958.2000.01691.x |
4. Hashimoto W, Nankai H, Sato N, Kawai S, and Murata K. (1999). Characterization of alpha-L-rhamnosidase of Bacillus sp. GL1 responsible for the complete depolymerization of gellan. Arch Biochem Biophys. 1999;368(1):56-60. DOI:10.1006/abbi.1999.1279 |
5. Manzanares P, van den Broeck HC, de Graaff LH, and Visser J. (2001). Purification and characterization of two different alpha-L-rhamnosidases, RhaA and RhaB, from Aspergillus aculeatus. Appl Environ Microbiol. 2001;67(5):2230-4. DOI:10.1128/AEM.67.5.2230-2234.2001 |
6. Koseki T, Mese Y, Nishibori N, Masaki K, Fujii T, Handa T, Yamane Y, Shiono Y, Murayama T, and Iefuji H. (2008). Characterization of an alpha-L-rhamnosidase from Aspergillus kawachii and its gene. Appl Microbiol Biotechnol. 2008;80(6):1007-13. DOI:10.1007/s00253-008-1599-7 |
7. Ichinose H, Fujimoto Z, and Kaneko S. (2013). Characterization of an α-L-Rhamnosidase from Streptomyces avermitilis. Biosci Biotechnol Biochem. 2013;77(1):213-6. DOI:10.1271/bbb.120735 |
8. Fujimoto Z, Jackson A, Michikawa M, Maehara T, Momma M, Henrissat B, Gilbert HJ, and Kaneko S. (2013). The structure of a Streptomyces avermitilis α-L-rhamnosidase reveals a novel carbohydrate-binding module CBM67 within the six-domain arrangement. J Biol Chem. 2013;288(17):12376-85. DOI:10.1074/jbc.M113.460097 |
9. Cui Z, Maruyama Y, Mikami B, Hashimoto W, and Murata K. (2007). Crystal structure of glycoside hydrolase family 78 alpha-L-Rhamnosidase from Bacillus sp. GL1. J Mol Biol. 2007;374(2):384-98. DOI:10.1016/j.jmb.2007.09.003 |
10. Bonanno JB, Almo SC, Bresnick A, Chance MR, Fiser A, Swaminathan S, Jiang J, Studier FW, Shapiro L, Lima CD, Gaasterland TM, Sali A, Bain K, Feil I, Gao X, Lorimer D, Ramos A, Sauder JM, Wasserman SR, Emtage S, D'Amico KL, and Burley SK. (2005). New York-Structural GenomiX Research Consortium (NYSGXRC): a large scale center for the protein structure initiative. J Struct Funct Genomics. 2005;6(2-3):225-32. DOI:10.1007/s10969-005-6827-0 |
11. Cantarel BL, Coutinho PM, Rancurel C, Bernard T, Lombard V, and Henrissat B. (2009). The Carbohydrate-Active EnZymes database (CAZy): an expert resource for Glycogenomics. Nucleic Acids Res. 2009;37(Database issue):D233-8. DOI:10.1093/nar/gkn663 |

12. Davies, G.J. and Sinnott, M.L. (2008) Sorting the diverse: the sequence-based classifications of carbohydrate-active enzymes. Biochem. J. (BJ Classic Paper, online only). DOI: 10.1042/BJ20080382