Glycoside Hydrolase Family 44

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• Author: ^^^Peter Reilly^^^
• Responsible Curator: ^^^Peter Reilly^^^

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Glycoside Hydrolase Family GH44
Clan	None specified, but Kitago et al. [1] and Nam et al. [2] suggest that it belongs to Clan GH-A.
Mechanism	Retaining
Active site residues	Catalytic proton donor/acceptor: Glu; catalytic nucleophile: Glu
CAZy DB link
http://www.cazy.org/GH44.html

Substrate specificities

Active on many substances, including cellooligosaccharides of DP4 and longer, carboxymethylcellulose, xylan, lichenan, Avicel (slightly), and xyloglucan, which appears to be a prime substrate [4,5].

Kinetics and Mechanism

Content is to be added here.

Catalytic Residues

Clostridium thermocellum endoglucanase: catalytic proton donor/acceptor: Glu186; catalytic nucleophile: Glu359 [1]. Protein from metagenomic library: catalytic proton donor/acceptor: Glu221; catalytic nucleophile: Glu393 [2].Clostridium acetobutylicum endoglucanase: catalytic proton donor/acceptor: Glu180; catalytic nucleophile: Glu352 [3].

Three-dimensional structures

The first three-dimensional structure was by Kitago et al. [1], who found a TIM-like barrel domain and a beta-sandwich domain in a Clostridium thermocellum endoglucanase. Similar structures were found by Nam et al. [2] in a protein from a metagenomic library and Warner et al. [3] in a Clostridium acetobutylicum endoglucanase. Ca and Zn ions are found as ligands [1].

Family Firsts

First stereochemistry determination

Kitago et al. [1] found that the enzyme acts by a retaining mechanism. They observed that a beta-anomer was preferentially formed during cyclohexaitol hydrolysis.

First catalytic nucleophile identification

Kitago et al. [1], by testing activity of the E359Q mutant.

First general acid/base residue identification

Kitago et al. [1], by testing activity of the E186Q mutant.

First 3-D structure

Kitago et al. [1] of an endoglucanase from Clostridium thermocellum. It had a resolution of 0.96 Å and allowed the identification of the catalytic residues and the mechanism.

References

1. Kitago Y, Karita S, Watanabe N, Kamiya M, Aizawa T, Sakka K, and Tanaka I. (2007). Crystal structure of Cel44A, a glycoside hydrolase family 44 endoglucanase from Clostridium thermocellum. J Biol Chem. 2007;282(49):35703-11. DOI:10.1074/jbc.M706835200 |
2. Nam KH, Kim SJ, and Hwang KY. (2009). Crystal structure of CelM2, a bifunctional glucanase-xylanase protein from a metagenome library. Biochem Biophys Res Commun. 2009;383(2):183-6. DOI:10.1016/j.bbrc.2009.03.149 |
3. Warner CD, Hoy JA, Shilling TC, Linnen MJ, Ginder ND, Ford CF, Honzatko RB, and Reilly PJ. (2010). Tertiary structure and characterization of a glycoside hydrolase family 44 endoglucanase from Clostridium acetobutylicum. Appl Environ Microbiol. 2010;76(1):338-46. DOI:10.1128/AEM.02026-09 |
4. Najmudin S, Guerreiro CI, Carvalho AL, Prates JA, Correia MA, Alves VD, Ferreira LM, Romão MJ, Gilbert HJ, Bolam DN, and Fontes CM. (2006). Xyloglucan is recognized by carbohydrate-binding modules that interact with beta-glucan chains. J Biol Chem. 2006;281(13):8815-28. DOI:10.1074/jbc.M510559200 |

5. Warner, CD, Go, RM, C. García-Salinas, C. Ford, and P. J. Reilly. 2010b. Kinetic characterization of a glycoside hydrolase family 44 endoglucanase from Ruminococcus flavefaciens FD-1. Submitted for publication.