Glycoside Hydrolase Family 82

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• Authors: ^^^Mirjam Czjzek^^^, ^^^Gurvan Michel^^^, and ^^^Etienne Rebuffet^^^
• Responsible Curator: ^^^Mirjam Czjzek^^^

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Glycoside Hydrolase Family GH82
Clan	none
Mechanism	inverting
Active site residues	known
CAZy DB link
https://www.cazy.org/GH82.html

Substrate specificities

At the moment, all characterised members of glycoside hydrolase family 82 enzymes cleave the β-1,4 galactosidic bond of the marine algal polysaccharide iota-carrageenan [1, 2, 3] yielding products of the neocarrabiose series.

Kinetics and Mechanism

Family 82 enzymes are inverting enzymes, as first shown by NMR [1] on the iota-carrageenase from Alteromonas fortis. The reaction seems to involve a chloride ion which through the Glu222 participated in the polarisation of the catalytic water molecule [2].

Catalytic Residues

From structural analysis the catalytic residues have been predicted to be two out of the three candidate amino acids Glu245, Asp247 or Glu310 in the A. fortis iota-carrageenase [4]. A study in 2010, which utilized site directed mutagenesis, has confirmed that Glu245 plays the role of the general acid residue in this inverting enzyme, while Asp247 is the general base activating the nucleophilic water molecule [2]. However, intriguingly the position of equivalent residues to Asp247 in other iota-carrageenase sequences are not strictly conserved [2, 3].

Three-dimensional structures

Figure 1: Movement of domain A induced by substrate binding. The β-helix is in blue , the domain A is in red and the domain B is in green .

A crystal structure has only been determined for the iota-carrageenase from A. fortis and the protein folds into a right-handed parallel β-helix of 10 complete turns with two additional C-terminal domains (A and B) [4]. The crystal structure of a product complex has shed light on the existence of domain movement of domain A that is closed around the oligo-carrageenan in the complexed form and open in the uncomplexed enzyme (Figure 1) [5]. Recent discovering of shorter sequences shows the existence of enzymes devoid of the C-terminal domain A or both domains A and B [2].

Family Firsts

First sequence identification and family creation

iota-carrageenase sequences have been first reported for enzymes from A. fortis and Z. galactanivorans [1].

First sterochemistry determination

GH82 enzymes are inverting as shown by NMR [1].

First general acid residue identification

Glu245 in the iota-carrageenase from Alteromonas fortis [2]

First general base residue identification

Asp247 in the iota-carrageenase from Alteromonas fortis [2].

First 3-D structure

iota-carrageenase from A. fortis [4]. The structure belongs to the β-helix fold (PDB 1h80 and PDB 1ktw).

References

1. Barbeyron T, Michel G, Potin P, Henrissat B, and Kloareg B. (2000). iota-Carrageenases constitute a novel family of glycoside hydrolases, unrelated to that of kappa-carrageenases. J Biol Chem. 2000;275(45):35499-505. DOI:10.1074/jbc.M003404200 |
2. Rebuffet E, Barbeyron T, Jeudy A, Jam M, Czjzek M, and Michel G. (2010). Identification of catalytic residues and mechanistic analysis of family GH82 iota-carrageenases. Biochemistry. 2010;49(35):7590-9. DOI:10.1021/bi1003475 |
3. Hatada Y, Mizuno M, Li Z, and Ohta Y. (2011). Hyper-production and characterization of the ι-carrageenase useful for ι-carrageenan oligosaccharide production from a deep-sea bacterium, Microbulbifer thermotolerans JAMB-A94T, and insight into the unusual catalytic mechanism. Mar Biotechnol (NY). 2011;13(3):411-22. DOI:10.1007/s10126-010-9312-0 |
4. Michel G, Chantalat L, Fanchon E, Henrissat B, Kloareg B, and Dideberg O. (2001). The iota-carrageenase of Alteromonas fortis. A beta-helix fold-containing enzyme for the degradation of a highly polyanionic polysaccharide. J Biol Chem. 2001;276(43):40202-9. DOI:10.1074/jbc.M100670200 |
5. Michel G, Helbert W, Kahn R, Dideberg O, and Kloareg B. (2003). The structural bases of the processive degradation of iota-carrageenan, a main cell wall polysaccharide of red algae. J Mol Biol. 2003;334(3):421-33. DOI:10.1016/j.jmb.2003.09.056 |