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Difference between revisions of "Glycoside Hydrolase Family 116"

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== Kinetics and Mechanism ==
 
== Kinetics and Mechanism ==
  
The enzymes of this family are ''[[retaining]]'' glycoside hydrolases and follow the classical [[Koshland double-displacement mechanism]] <cite>Koshland</cite>. The stereochemistry of hydrolysis has been demonstrated by <sup>1</sup>H-<sup>13</sup>C NMR spectroscopy analysis of the interglycosidic linkage of disaccharides formed by the [[transglycosylation]] action of SSO1353 with 4NP-β-Xyl <cite>PMID20427274</cite>.
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The enzymes of this family are ''[[retaining]]'' glycoside hydrolases and follow the classical [[Koshland double-displacement mechanism]] <cite>Koshland</cite>. The stereochemistry of hydrolysis has been demonstrated by <sup>1</sup>H-<sup>13</sup>C NMR spectroscopy analysis of the interglycosidic linkage of disaccharides formed by the transglycosylation action of SSO1353 with 4NP-β-Xyl <cite>PMID20427274</cite>.
  
  

Revision as of 07:36, 7 July 2010

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Glycoside Hydrolase Family GH116
Clan none
Mechanism retaining
Active site residues known
CAZy DB link
http://www.cazy.org/GH116.html


Substrate specificities

This family of glycoside hydrolases was recently discovered characterising a new β-glycosidase from the hyperthermophilic archaeon Sulfolobus solfataricus [1] and contains acid β-glucosidase (EC 3.2.1.45), β-glucosidase (EC 3.2.1.21) and β-xylosidase (EC 3.2.1.37) activities from the three domains of life. The β-glycosidase from S. solfataricus (SSO1353) is specific for the gluco- and xylosides β-bound to hydrophobic groups that are hydrolyzed by following a retaining reaction mechanism. SSO1353 is distantly related to the human non-lysosomal bile acid β-glucosidase GBA2, also known as glucocerebrosidase, involved in the catabolism of glucosylceramide, which is then converted to sphingomyelin [2]. SSO1353 has substrate specificity and inhibitor sensitivity slightly different from those of GBA2. In fact, the archaeal enzyme can hydrolyze both aryl β-gluco and β-xylosides and it is inhibited by both N-butyldeoxynojirimycin (NB-DNJ) and conduritol β-epoxide (CBE) [1]. Instead, GBA2 is inactive on methylumbellyferyl-β-D-xylopyranoside and is relatively insensitive to CBE [2].


Kinetics and Mechanism

The enzymes of this family are retaining glycoside hydrolases and follow the classical Koshland double-displacement mechanism [3]. The stereochemistry of hydrolysis has been demonstrated by 1H-13C NMR spectroscopy analysis of the interglycosidic linkage of disaccharides formed by the transglycosylation action of SSO1353 with 4NP-β-Xyl [1].


Catalytic Residues

The catalytic residues were identified in the S. solfataricus β-glycosidase [1]. The catalytic nucleophile was identified as Glu335 through trapping of the 2- deoxy-2-fluoroglucosyl-enzyme intermediate and MS/MS analysis. The general acid/base catalyst role was assigned to Asp462 through mechanistic analysis of a mutant at that position, which included azide rescue experiments.


Three-dimensional structures

Unkwown


Family Firsts

First stereochemistry determination
S. solfataricus β-glycosidase by NMR [1].
First catalytic nucleophile identification
S. solfataricus β-glycosidase by 2-deoxy-2-fluoroglucose labeling [1].
First general acid/base residue identification
S. solfataricus β-glycosidase by azide rescue with mutant [1].
First 3-D structure


References

  1. Cobucci-Ponzano B, Aurilia V, Riccio G, Henrissat B, Coutinho PM, Strazzulli A, Padula A, Corsaro MM, Pieretti G, Pocsfalvi G, Fiume I, Cannio R, Rossi M, and Moracci M. (2010). A new archaeal beta-glycosidase from Sulfolobus solfataricus: seeding a novel retaining beta-glycan-specific glycoside hydrolase family along with the human non-lysosomal glucosylceramidase GBA2. J Biol Chem. 2010;285(27):20691-703. DOI:10.1074/jbc.M109.086470 | PubMed ID:20427274 [PMID20427274]
  2. Boot RG, Verhoek M, Donker-Koopman W, Strijland A, van Marle J, Overkleeft HS, Wennekes T, and Aerts JM. (2007). Identification of the non-lysosomal glucosylceramidase as beta-glucosidase 2. J Biol Chem. 2007;282(2):1305-12. DOI:10.1074/jbc.M610544200 | PubMed ID:17105727 [PMID17105727]
  3. Koshland DE Jr: Stereochemistry and the mechanism of enzyme reactions. Biol Rev 1953, 28:416-436.

    [Koshland]

All Medline abstracts: PubMed