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Difference between revisions of "Glycoside Hydrolase Family 92"
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== Substrate specificities == | == Substrate specificities == | ||
| − | The [[glycoside hydrolases]] of GH92 are [[exo]]-acting α-mannosidases. The first reported enzyme activity from this family was an α-1,2-mannosidase from ''Microbacterium'' sp. M-90 | + | The [[glycoside hydrolases]] of GH92 are [[exo]]-acting α-mannosidases. The first reported enzyme activity from this family was an α-1,2-mannosidase from ''Microbacterium'' sp. M-90 <cite>#1</cite>. Recently the characterization of 22 GH92 enzymes from ''Bacteroides thetaiotaomicron'' confirmed an [[exo]]-mode of action with α-1,2-mannosidase, α-1,3-mannosidase, α-1,4-mannosidase and α-1,6-mannosidase activities were detected <cite>#2</cite>. |
== Kinetics and Mechanism == | == Kinetics and Mechanism == | ||
| − | <sup>1</sup>H | + | <sup>1</sup>H NMR studies on three GH92s that displayed α-1,2-, α-1,3- and α-1,4-mannosidase activities all generated β-mannose indicating that these enzymes catalyse glycosidic bond hydrolysis through a single displacement mechanism leading to [[inverting|inversion]] of anomeric configuration <cite>#2</cite>. GH92 enzymes are Ca<sup>2+</sup>-dependent α-mannosidases. The requirement for the metal ion is currently restricted to only three GH families all of which are [[exo]]-α-mannosidases: [[GH38]], [[GH47]] and GH92. Mechanistically this may indicate that the lack of distorting binding energy provided by the -2 or +1 subsites impose a requirement for conformational flexibility at the -1 subsite (recognition of the ground state and the transition state conformations), which is achieved by a metal ion interaction with O2 and O3. Three inhibitors bound to the α-1,2-mannosidase Bt3990 in approximate <sup>1</sup>''S''<sub>5</sub>/''B''<sub>2,5</sub> and <sup>1,4</sup>''B''/<sup>1</sup>''S''<sub>5</sub> conformations indicating that catalysis is proceeds via a ''B''<sub>2,5</sub> transition state. |
== Catalytic Residues == | == Catalytic Residues == | ||
| Line 40: | Line 40: | ||
== Family Firsts == | == Family Firsts == | ||
| − | ;First sterochemistry determination: <sup>1</sup>H | + | ;First sterochemistry determination: <sup>1</sup>H NMR showed three GH92s generate β-mannose and thus these α-mannosidases are [[inverting]] enzymes <cite>#2</cite>. |
;First [[general acid]] identification: Based on mutagenesis and 3D structural information the conserved catalytic acid has been identified <cite>#2</cite>. | ;First [[general acid]] identification: Based on mutagenesis and 3D structural information the conserved catalytic acid has been identified <cite>#2</cite>. | ||
;First [[general base]] residue identification: Based on mutagenesis and 3D structural information a pair of likely catalytic bases were identified. As one of these residues is invariant this is the proposed catalytic base <cite>#2</cite>. | ;First [[general base]] residue identification: Based on mutagenesis and 3D structural information a pair of likely catalytic bases were identified. As one of these residues is invariant this is the proposed catalytic base <cite>#2</cite>. | ||
Revision as of 01:46, 12 January 2012
This page has been approved by the Responsible Curator as essentially complete. CAZypedia is a living document, so further improvement of this page is still possible. If you would like to suggest an addition or correction, please contact the page's Responsible Curator directly by e-mail.
- Author: ^^^Harry Gilbert^^^
- Responsible Curator: ^^^Harry Gilbert^^^
| Glycoside Hydrolase Family GH92 | |
| Clan | GH-x |
| Mechanism | inverting |
| Active site residues | known/not known |
| CAZy DB link | |
| http://www.cazy.org/GH92.html | |
Substrate specificities
The glycoside hydrolases of GH92 are exo-acting α-mannosidases. The first reported enzyme activity from this family was an α-1,2-mannosidase from Microbacterium sp. M-90 [1]. Recently the characterization of 22 GH92 enzymes from Bacteroides thetaiotaomicron confirmed an exo-mode of action with α-1,2-mannosidase, α-1,3-mannosidase, α-1,4-mannosidase and α-1,6-mannosidase activities were detected [2].
Kinetics and Mechanism
1H NMR studies on three GH92s that displayed α-1,2-, α-1,3- and α-1,4-mannosidase activities all generated β-mannose indicating that these enzymes catalyse glycosidic bond hydrolysis through a single displacement mechanism leading to inversion of anomeric configuration [2]. GH92 enzymes are Ca2+-dependent α-mannosidases. The requirement for the metal ion is currently restricted to only three GH families all of which are exo-α-mannosidases: GH38, GH47 and GH92. Mechanistically this may indicate that the lack of distorting binding energy provided by the -2 or +1 subsites impose a requirement for conformational flexibility at the -1 subsite (recognition of the ground state and the transition state conformations), which is achieved by a metal ion interaction with O2 and O3. Three inhibitors bound to the α-1,2-mannosidase Bt3990 in approximate 1S5/B2,5 and 1,4B/1S5 conformations indicating that catalysis is proceeds via a B2,5 transition state.
Catalytic Residues
Based on 3D structural data on the α-1,2-mannosidase Bt3990, Glu533 is the predicted general acid. This view is supported by an inactive mutant of this residue, and the conservation of the glutamate throughout the GH92 family [2]. The general base, in common with many inverting glycoside hydrolases, is more difficult to identify. Asp644 and Asp642 both lie in the canonical position expected for a general base in an inverting enzyme. Mutants of both residues inactivte the enzyme, however, while Asp644 is invariant, Asp642 can be an Asn or Asp in GH92 members [2]. It appears that Asp644 is the likely catalytic general base.
Three-dimensional structures
GH92 enzymes display a two domain structure. The small N-terminal domain is a beta-sandwich and the large C-terminal domain adopts a adorned (alpha/alpha)6 barrel fold. Amino acids in the active site of the enzyme, a shallow pocket, are contributed by both the N- and C-terminal domains [2].
Family Firsts
- First sterochemistry determination
- 1H NMR showed three GH92s generate β-mannose and thus these α-mannosidases are inverting enzymes [2].
- First general acid identification
- Based on mutagenesis and 3D structural information the conserved catalytic acid has been identified [2].
- First general base residue identification
- Based on mutagenesis and 3D structural information a pair of likely catalytic bases were identified. As one of these residues is invariant this is the proposed catalytic base [2].
- First 3-D structure
- The 3D structure reveals two domains; an N-terminal β-sandwich domain and a C-terminal adorned (α/α)6 barrel. Both domains contribute residues to the active site [2].
References
- Maruyama Y, Nakajima T, and Ichishima E. (1994). A 1,2-alpha-D-mannosidase from a Bacillus sp.: purification, characterization, and mode of action. Carbohydr Res. 1994;251:89-98. DOI:10.1016/0008-6215(94)84278-7 |
- Zhu Y, Suits MD, Thompson AJ, Chavan S, Dinev Z, Dumon C, Smith N, Moremen KW, Xiang Y, Siriwardena A, Williams SJ, Gilbert HJ, and Davies GJ. (2010). Mechanistic insights into a Ca2+-dependent family of alpha-mannosidases in a human gut symbiont. Nat Chem Biol. 2010;6(2):125-32. DOI:10.1038/nchembio.278 |