File:Hidaka.jpg

2009-07-28T03:54:22Z

Masafumi Hidaka:

Glycoside Hydrolase Family 94

2009-07-28T03:44:26Z

Masafumi Hidaka: /* References */

* [[Author]]: [[User:Masafumi_Hidaka|Masafumi Hidaka]]
* [[Responsible Curator]]: [[User:ShinyaFushinobu|Shinya Fushinobu]]
----

<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Glycoside Hydrolase Family 94'''
|-
|'''Clan'''
|none (similar to GH-L)
|-
|'''Mechanism'''
|inverting
|-
|'''Active site residues'''
|known
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |http://www.cazy.org/fam/GH94.html
|}
</div>

== Substrate specificities ==
This family exclusively contains [[Phosphorylases]] that cleave β-glycosidic bond. The substrate specificities found in GH94 are: cellobiose (Glc-β1,4-Glc) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.20 2.4.1.20]), cellodextrin ((Glc-β1,4-)<sub>n-1</sub>Glc; n ≥ 3) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.29 2.4.1.29]), and N,N’-diacetyl chitobiose (GlcNAc-β1,4-GlcNAc) phosphorylase. Moreover, a phosphorylase domain belonging to this family is found in cyclic β-1,2-glucan synthase, a modular protein that also contains a glycosyltransferase domain from [[GlycosylTransferase Family 84]] <cite>REF7</cite>. The GH94 domain is thought to phosphorolyze protein-bound β-1,2-glucans synthesized from UDP-glucose by the GT84 domain.<BR>

GH94 enzymes were initially classified in [[GlycosylTransferase Family 36]] because none of them show hydrolytic activity. However because of the evolutionary, structural and mechanistic relatedness with clan GH-L glycoside hydrolases, the family was re-assigned to family GH94 <cite>REF1</cite>.

== Kinetics and Mechanism ==
Phosphorolysis by GH94 enzymes proceeds with inversion of anomeric configuration, as first shown by Sih and McBee <cite>REF5</cite> on cellobiose phosphorylase from ''Clostridium thermocellum'', i.e. cellobiose (Glc-β1,4-Glc) + Pi ↔ α-glucose 1-phosphate + glucose. Considering the topology of the active site structure, the reaction mechanism for inverting phosphorylase is proposed to be similar to that for inverting GH <cite>REF1</cite>. With the aid of general acid residue, the enzymatic phosphorolysis begins with direct nucleophilic attack by phosphate on the anomeric C-1 carbon, instead of the water molecule activated by a general base residue in inverting GH reaction.

== Catalytic Residues ==
The catalytic residue was firstly estimated by superimposing the active site structure of chitobiose phosphorylase from ''Vibrio proteolyticus'' with a [[Glycoside Hydrolase Family 15]] enzyme, glucoamylase from ''Thermoanaerobacterium thermosaccharolyticum'' <cite>REF1</cite>. Considering the similarities of the active site structure, Asp492 was estimated as the general acid residue. D492A/N mutants of this enzyme showed no detectable activity. General base residue is not required for the reaction of glycoside hydrolase-like inverting phosphorylases.

== Three-dimensional structures ==
The first solved 3-D structure was chitobiose phosphorylase from ''Vibrio proteolyticus'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7V 1V7V], [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7W 1V7W],
[http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7X 1V7X]) <cite>REF1</cite>. The enzyme has a (α/α)<sub>6</sub> barrel fold that is remarkably similar to clan GH-L. The position of the catalytic general acid is superimposable with Clan GH-L. It should be noted that GH94 enzymes act on β-bonds, whereas clan GH-L enzymes ([[Glycoside Hydrolase Family 15]] and [[Glycoside Hydrolase Family 65]]) act on α-bonds.

== Family Firsts ==
;First sterochemistry determination:
Cellobiose phosphorylase from ''Clostridium thermocellum'' <cite>REF5</cite>
;First gene cloning:
Cellobiose phosphorylase and a cellodextrin phosphorylase from ''Clostridium stercorarium'' <cite>REF6</cite>
;First catalytic nucleophile identification:
The inverting phosphorolytic reaction does not require catalytic general base residue, but inorganic phosphate act as a nucleophile.
;First general acid residue identification:
''Vibrio proteolyticus'' chitobiose phosphorylase by kinetic studies with mutants <cite>REF1</cite>
;First 3-D structure:
''Vibrio proteolyticus'' chitobiose phosphorylase <cite>REF1</cite>.

== References ==
<biblio>
#REF1 pmid=15274915
#REF5 Sih CJ, and McBee RH. ''A cellobiose phosphorylase in Clostridium thermocellum.'' Proc Montana Acad Sci 1955, 15, 21-22.

#REF6 pmid=9249035
#REF7 pmid=17921247

</biblio>

[[Category:Glycoside Hydrolase Families]]

Glycoside Hydrolase Family 94

2009-07-16T00:30:56Z

Masafumi Hidaka: /* Three-dimensional structures */

* [[Author]]: [[User:Masafumi_Hidaka|Masafumi Hidaka]]
* [[Responsible Curator]]: [[User:ShinyaFushinobu|Shinya Fushinobu]]
----

<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Glycoside Hydrolase Family 94'''
|-
|'''Clan'''
|none (similar to GH-L)
|-
|'''Mechanism'''
|inverting
|-
|'''Active site residues'''
|known
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |http://www.cazy.org/fam/GH94.html
|}
</div>

== Substrate specificities ==
This family contains phosphorolytic enzymes (usually named using a combination of “the substrate” and “phosphorylase”) that cleave beta glycosidic bond. The substrate specificities found in GH94 are: cellobiose (Glc-β1,4-Glc) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.20 2.4.1.20]), cellodextrin ((Glc-β1,4-)<sub>n-1</sub>Glc; n ≥ 3) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.29 2.4.1.29]), and N,N’-diacetyl chitobiose (GlcNAc-β1,4-GlcNAc) phosphorylase. Moreover, a phosphorylase domain belonging to this family is found in cyclic β-1,2-glucan synthase, a modular protein that also contains a glycosyltransferase domain from [[GlycosylTransferase Family 84]] <cite>REF7</cite>. The GH94 domain is thought to phosphorolyze protein-bound β-1,2-glucans synthesized from UDP-glucose by the GT84 domain.<BR> [[Phosphorylases]] catalyze the phosphorolysis of glycosidic bonds to generate glycosyl-phosphate. The reaction is reversible due to the energy of the glycosyl-phosphate bond. Therefore, phosphorylases are categorized as “transferase” among enzyme nomenclature (EC 2.4.1.-). Together with the fact that the GH94 enzymes did not show hydrolytic activity, GH94 enzymes were initially classified in [[GlycosylTransferase Family 36]]. However because of the evolutionary, structural and mechanistic relatedness of GH94 phosphorylases with clan GH-L glycoside hydrolases, the family was re-assigned to family GH94 <cite>REF1</cite>.

== Kinetics and Mechanism ==
Phosphorolysis by GH94 enzymes proceeds with inversion of anomeric configuration, as first shown by Sih and McBee <cite>REF5</cite> on cellobiose phosphorylase from ''Clostridium thermocellum'', i.e. cellobiose (Glc-β1,4-Glc) + Pi ↔ α-glucose 1-phosphate + glucose. Considering the topology of the active site structure, the reaction mechanism for inverting phosphorylase is proposed to be similar to that for inverting GH <cite>REF1</cite>. With the aid of general acid residue, the enzymatic phosphorolysis begins with direct nucleophilic attack by phosphate on the anomeric C-1 carbon, instead of the water molecule activated by a general base residue in inverting GH reaction.

== Catalytic Residues ==
The catalytic residue was firstly estimated by superimposing the active site structure of chitobiose phosphorylase from ''Vibrio proteolyticus'' with a [[Glycoside Hydrolase Family 15]] enzyme, glucoamylase from ''Thermoanaerobacterium thermosaccharolyticum'' <cite>REF1</cite>. Considering the similarities of the active site structure, Asp492 was estimated as the general acid residue. D492A/N mutants of this enzyme showed no detectable activity. General base residue is not required for the reaction of glycoside hydrolase-like inverting phosphorylases.

== Three-dimensional structures ==
The first solved 3-D structure was chitobiose phosphorylase from ''Vibrio proteolyticus'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7V 1V7V], [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7W 1V7W],
[http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7X 1V7X]) <cite>REF1</cite>. The enzyme has a (α/α)<sub>6</sub> barrel fold that is remarkably similar to clan GH-L. The position of the catalytic general acid is superimposable with Clan GH-L. It should be noted that GH94 enzymes act on β-bonds, whereas clan GH-L enzymes ([[Glycoside Hydrolase Family 15]] and [[Glycoside Hydrolase Family 65]]) act on α-bonds.

== Family Firsts ==
;First sterochemistry determination:
Cellobiose phosphorylase from ''Clostridium thermocellum'' <cite>REF5</cite>
;First gene cloning:
Cellobiose phosphorylase and a cellodextrin phosphorylase from ''Clostridium stercorarium'' <cite>REF6</cite>
;First catalytic nucleophile identification:
The inverting phosphorolytic reaction does not require catalytic general base residue, but inorganic phosphate act as a nucleophile.
;First general acid residue identification:
''Vibrio proteolyticus'' chitobiose phosphorylase by kinetic studies with mutants <cite>REF1</cite>
;First 3-D structure:
''Vibrio proteolyticus'' chitobiose phosphorylase <cite>REF1</cite>.

== References ==
<biblio>
#REF1 pmid=15274915
#REF2 pmid=14756551
#REF3 pmid=11587643
#REF4 pmid=19124470
#REF5 Sih CJ, and McBee RH. ''A cellobiose phosphorylase in Clostridium thermocellum.'' Proc Montana Acad Sci 1955, 15, 21-22.

#REF6 pmid=9249035
#REF7 pmid=17921247

</biblio>

[[Category:Glycoside Hydrolase Families]]

Glycoside Hydrolase Family 94

2009-07-16T00:28:17Z

Masafumi Hidaka: /* Substrate specificities */

* [[Author]]: [[User:Masafumi_Hidaka|Masafumi Hidaka]]
* [[Responsible Curator]]: [[User:ShinyaFushinobu|Shinya Fushinobu]]
----

<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Glycoside Hydrolase Family 94'''
|-
|'''Clan'''
|none (similar to GH-L)
|-
|'''Mechanism'''
|inverting
|-
|'''Active site residues'''
|known
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |http://www.cazy.org/fam/GH94.html
|}
</div>

== Substrate specificities ==
This family contains phosphorolytic enzymes (usually named using a combination of “the substrate” and “phosphorylase”) that cleave beta glycosidic bond. The substrate specificities found in GH94 are: cellobiose (Glc-β1,4-Glc) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.20 2.4.1.20]), cellodextrin ((Glc-β1,4-)<sub>n-1</sub>Glc; n ≥ 3) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.29 2.4.1.29]), and N,N’-diacetyl chitobiose (GlcNAc-β1,4-GlcNAc) phosphorylase. Moreover, a phosphorylase domain belonging to this family is found in cyclic β-1,2-glucan synthase, a modular protein that also contains a glycosyltransferase domain from [[GlycosylTransferase Family 84]] <cite>REF7</cite>. The GH94 domain is thought to phosphorolyze protein-bound β-1,2-glucans synthesized from UDP-glucose by the GT84 domain.<BR> [[Phosphorylases]] catalyze the phosphorolysis of glycosidic bonds to generate glycosyl-phosphate. The reaction is reversible due to the energy of the glycosyl-phosphate bond. Therefore, phosphorylases are categorized as “transferase” among enzyme nomenclature (EC 2.4.1.-). Together with the fact that the GH94 enzymes did not show hydrolytic activity, GH94 enzymes were initially classified in [[GlycosylTransferase Family 36]]. However because of the evolutionary, structural and mechanistic relatedness of GH94 phosphorylases with clan GH-L glycoside hydrolases, the family was re-assigned to family GH94 <cite>REF1</cite>.

== Kinetics and Mechanism ==
Phosphorolysis by GH94 enzymes proceeds with inversion of anomeric configuration, as first shown by Sih and McBee <cite>REF5</cite> on cellobiose phosphorylase from ''Clostridium thermocellum'', i.e. cellobiose (Glc-β1,4-Glc) + Pi ↔ α-glucose 1-phosphate + glucose. Considering the topology of the active site structure, the reaction mechanism for inverting phosphorylase is proposed to be similar to that for inverting GH <cite>REF1</cite>. With the aid of general acid residue, the enzymatic phosphorolysis begins with direct nucleophilic attack by phosphate on the anomeric C-1 carbon, instead of the water molecule activated by a general base residue in inverting GH reaction.

== Catalytic Residues ==
The catalytic residue was firstly estimated by superimposing the active site structure of chitobiose phosphorylase from ''Vibrio proteolyticus'' with a [[Glycoside Hydrolase Family 15]] enzyme, glucoamylase from ''Thermoanaerobacterium thermosaccharolyticum'' <cite>REF1</cite>. Considering the similarities of the active site structure, Asp492 was estimated as the general acid residue. D492A/N mutants of this enzyme showed no detectable activity. General base residue is not required for the reaction of glycoside hydrolase-like inverting phosphorylases.

== Three-dimensional structures ==
The first solved 3-D structure was chitobiose phosphorylase from ''Vibrio proteolyticus'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7V 1V7V], [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7W 1V7W],
[http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7X 1V7X]) <cite>REF1</cite>. The enzyme has a (α/α)<sub>6</sub> barrel fold that is remarkably similar to clan GH-L. The position of the catalytic general acid is superimposable with Clan GH-L. It should be noted that GH94 enzymes act on β-bonds, whereas clan GH-L enzymes (GH15 and GH65) act on α-bonds.

== Family Firsts ==
;First sterochemistry determination:
Cellobiose phosphorylase from ''Clostridium thermocellum'' <cite>REF5</cite>
;First gene cloning:
Cellobiose phosphorylase and a cellodextrin phosphorylase from ''Clostridium stercorarium'' <cite>REF6</cite>
;First catalytic nucleophile identification:
The inverting phosphorolytic reaction does not require catalytic general base residue, but inorganic phosphate act as a nucleophile.
;First general acid residue identification:
''Vibrio proteolyticus'' chitobiose phosphorylase by kinetic studies with mutants <cite>REF1</cite>
;First 3-D structure:
''Vibrio proteolyticus'' chitobiose phosphorylase <cite>REF1</cite>.

== References ==
<biblio>
#REF1 pmid=15274915
#REF2 pmid=14756551
#REF3 pmid=11587643
#REF4 pmid=19124470
#REF5 Sih CJ, and McBee RH. ''A cellobiose phosphorylase in Clostridium thermocellum.'' Proc Montana Acad Sci 1955, 15, 21-22.

#REF6 pmid=9249035
#REF7 pmid=17921247

</biblio>

[[Category:Glycoside Hydrolase Families]]

User:Masafumi Hidaka

2009-07-14T23:21:55Z

Masafumi Hidaka:

'''Masafumi Hidaka''' is a postdoctoral fellow in [http://enzyme13.bt.a.u-tokyo.ac.jp the Laboratory of Enzymology at Department of Biotechnology, The University of Tokyo]. He received his Ph.D. from the Graduate School of Agricultural and Life Sciences, The University of Tokyo in 2005. His research interests are the structural insights into mechanism of enzymatic reaction.
[[Category:Contributors|Hidaka, Masafumi]]

User:Masafumi Hidaka

2009-07-14T23:20:33Z

Masafumi Hidaka:

Glycoside Hydrolase Family 94

2009-07-14T23:18:05Z

Masafumi Hidaka: /* Three-dimensional structures */

* [[Author]]: [[User:Masafumi_Hidaka|Masafumi Hidaka]]
* [[Responsible Curator]]: [[User:ShinyaFushinobu|Shinya Fushinobu]]
----

<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Glycoside Hydrolase Family 94'''
|-
|'''Clan'''
|none (similar to GH-L)
|-
|'''Mechanism'''
|inverting
|-
|'''Active site residues'''
|known
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |http://www.cazy.org/fam/GH94.html
|}
</div>

== Substrate specificities ==
This family contains phosphorolytic enzymes (usually named using a combination of “the substrate” and “phosphorylase”) that cleave beta glycosidic bond. The substrate specificities found in GH94 are: cellobiose (Glc-β1,4-Glc) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.20 2.4.1.20]), cellodextrin ((Glc-β1,4-)<sub>n-1</sub>Glc; n ≥ 3) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.29 2.4.1.29]), and N,N’-diacetyl chitobiose (GlcNAc-β1,4-GlcNAc) phosphorylase. Moreover, a phosphorylase domain belonging to this family is found in cyclic β-1,2-glucan synthase (EC 2.4.1.-) along with a [[GlycosylTransferase Family 84]] domain <cite>REF7</cite>. This domain is thought to phosphorolyze protein-bound β-1,2-glucans.<BR> '''Phosphorylases''' catalyze the phosphorolysis of glycosidic bonds to generate glycosyl-phosphate. The reaction is reversible due to the energy of the glycosyl-phosphate bond. Therefore, phosphorylases are categorized as “transferase” among enzyme nomenclature (EC 2.4.1.-). Together with the fact that the GH94 enzymes did not show hydrolytic activity, GH94 enzymes were initially classified in [[GlycosylTransferase Family 36]]. However because of the evolutionary, structural and mechanistic relatedness of GH94 phosphorylases with clan GH-L glycoside hydrolases, the family was re-assigned to family GH94 <cite>REF1</cite>.

== Kinetics and Mechanism ==
Phosphorolysis by GH94 enzymes proceeds with inversion of anomeric configuration, as first shown by Sih and McBee <cite>REF5</cite> on cellobiose phosphorylase from ''Clostridium thermocellum'', i.e. cellobiose (Glc-β1,4-Glc) + Pi ↔ α-glucose 1-phosphate + glucose. Considering the topology of the active site structure, the reaction mechanism for inverting phosphorylase is proposed to be similar to that for inverting GH <cite>REF1</cite>. With the aid of general acid residue, the enzymatic phosphorolysis begins with direct nucleophilic attack by phosphate on the anomeric C-1 carbon, instead of the water molecule activated by a general base residue in inverting GH reaction.

== Catalytic Residues ==
The catalytic residue was firstly estimated by superimposing the active site structure of chitobiose phosphorylase from ''Vibrio proteolyticus'' with a [[Glycoside Hydrolase Family 15]] enzyme, glucoamylase from ''Thermoanaerobacterium thermosaccharolyticum'' <cite>REF1</cite>. Considering the similarities of the active site structure, Asp492 was estimated as the general acid residue. D492A/N mutants of this enzyme showed no detectable activity. General base residue is not required for the reaction of glycoside hydrolase-like inverting phosphorylases.

== Three-dimensional structures ==
The first solved 3-D structure was chitobiose phosphorylase from ''Vibrio proteolyticus'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7V 1V7V], [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7W 1V7W],
[http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7X 1V7X]) <cite>REF1</cite>. The enzyme has a (α/α)<sub>6</sub> barrel fold that is remarkably similar to clan GH-L. The position of the catalytic general acid is superimposable with Clan GH-L. It should be noted that GH94 enzymes act on β-bonds, whereas clan GH-L enzymes (GH15 and GH65) act on α-bonds.

== Family Firsts ==
;First sterochemistry determination:
Cellobiose phosphorylase from ''Clostridium thermocellum'' <cite>REF5</cite>
;First gene cloning:
Cellobiose phosphorylase and a cellodextrin phosphorylase from ''Clostridium stercorarium'' <cite>REF6</cite>
;First catalytic nucleophile identification:
The inverting phosphorolytic reaction does not require catalytic general base residue, but inorganic phosphate act as a nucleophile.
;First general acid residue identification:
''Vibrio proteolyticus'' chitobiose phosphorylase by kinetic studies with mutants <cite>REF1</cite>
;First 3-D structure:
''Vibrio proteolyticus'' chitobiose phosphorylase <cite>REF1</cite>.

== References ==
<biblio>
#REF1 pmid=15274915
#REF2 pmid=14756551
#REF3 pmid=11587643
#REF4 pmid=19124470
#REF5 Sih CJ, and McBee RH. ''A cellobiose phosphorylase in Clostridium thermocellum.'' Proc Montana Acad Sci 1955, 15, 21-22.

#REF6 pmid=9249035
#REF7 pmid=17921247

</biblio>

[[Category:Glycoside Hydrolase Families]]

Glycoside Hydrolase Family 94

2009-07-14T15:02:08Z

Masafumi Hidaka: /* Three-dimensional structures */

* [[Author]]: [[User:Masafumi_Hidaka|Masafumi Hidaka]]
* [[Responsible Curator]]: [[User:ShinyaFushinobu|Shinya Fushinobu]]
----

<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Glycoside Hydrolase Family 94'''
|-
|'''Clan'''
|none (similar to GH-L)
|-
|'''Mechanism'''
|inverting
|-
|'''Active site residues'''
|known
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |http://www.cazy.org/fam/GH94.html
|}
</div>

== Substrate specificities ==
This family contains phosphorolytic enzymes (usually named using a combination of “the substrate” and “phosphorylase”) that cleave beta glycosidic bond. The substrate specificities found in GH94 are: cellobiose (Glc-β1,4-Glc) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.20 2.4.1.20]), cellodextrin ((Glc-β1,4-)<sub>n-1</sub>Glc; n ≥ 3) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.29 2.4.1.29]), and N,N’-diacetyl chitobiose (GlcNAc-β1,4-GlcNAc) phosphorylase. Moreover, a phosphorylase domain belonging to this family is found in cyclic β-1,2-glucan synthase (EC 2.4.1.-) along with a [[GlycosylTransferase Family 84]] domain <cite>REF7</cite>. This domain is thought to phosphorolyze protein-bound β-1,2-glucans.<BR> '''Phosphorylases''' catalyze the phosphorolysis of glycosidic bonds to generate glycosyl-phosphate. The reaction is reversible due to the energy of the glycosyl-phosphate bond. Therefore, phosphorylases are categorized as “transferase” among enzyme nomenclature (EC 2.4.1.-). Together with the fact that the GH94 enzymes did not show hydrolytic activity, GH94 enzymes were initially classified in [[GlycosylTransferase Family 36]]. However because of the evolutionary, structural and mechanistic relatedness of GH94 phosphorylases with clan GH-L glycoside hydrolases, the family was re-assigned to family GH94 <cite>REF1</cite>.

== Kinetics and Mechanism ==
Phosphorolysis by GH94 enzymes proceeds with inversion of anomeric configuration, as first shown by Sih and McBee <cite>REF5</cite> on cellobiose phosphorylase from ''Clostridium thermocellum'', i.e. cellobiose (Glc-β1,4-Glc) + Pi ↔ α-glucose 1-phosphate + glucose. Considering the topology of the active site structure, the reaction mechanism for inverting phosphorylase is proposed to be similar to that for inverting GH <cite>REF1</cite>. With the aid of general acid residue, the enzymatic phosphorolysis begins with direct nucleophilic attack by phosphate on the anomeric C-1 carbon, instead of the water molecule activated by a general base residue in inverting GH reaction.

== Catalytic Residues ==
The catalytic residue was firstly estimated by superimposing the active site structure of chitobiose phosphorylase from ''Vibrio proteolyticus'' with a [[Glycoside Hydrolase Family 15]] enzyme, glucoamylase from ''Thermoanaerobacterium thermosaccharolyticum'' <cite>REF1</cite>. Considering the similarities of the active site structure, Asp492 was estimated as the general acid residue. D492A/N mutants of this enzyme showed no detectable activity. General base residue is not required for the reaction of glycoside hydrolase-like inverting phosphorylases.

== Three-dimensional structures ==
The first solved 3-D structure was chitobiose phosphorylase from ''Vibrio proteolyticus'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7V 1V7V], [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7W 1V7W],
[http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7X 1V7X]) <cite>REF1</cite>. The enzyme has a (α/α)<sub>6</sub> barrel fold that is remarkably similar to clan GH-L. The position of the catalytic general acid is superimposable with Clan GH-L. It should be noted that GH94 enzymes act on β-bonds, whereas clan GH-L enzymes (GH15 and GH65) act on α-bonds. <BR> Today, phosphorylases are categorized based on the evolutionary origins. GH type phosphorylases are classified in [[Glycoside Hydrolase Family 13]], [[Glycoside Hydrolase Family 65]], GH94, and [[Glycoside Hydrolase Family 112]]. GH13 sucrose phosphorylase from ''Bifidobacterium adolescentis'' has a TIM barrel fold catalytic domain like other GH13 hydorolytic enzymes (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1R7A 1R7A]) <cite>REF2</cite>. GH65 maltose phorphorylase from ''Lactobacillus brevis'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1H54 1H54]) <cite>REF3</cite> and GH94 enzymes share clan GH-L like (α/α)<sub>6</sub> barrel fold domain. GH112 galacto-''N''-biose/lacto-''N''-biose I phosphorylase (β-1,3-D-galactosyl-D-hexososamine phosphorylase) from ''Bifidobacterium longum'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUS 2ZUS], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUT 2ZUT], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUU 2ZUU], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUV 2ZUV], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUW 2ZUW] ), which catalyzes phosphorolysis of β-galactosidic bond, has a TIM barrel fold domain similar with that of GH42 β-galactosidase, hydrolase for β-galactosidic bond <cite>REF4</cite>. GT-type phosphorylases are classified in GT4 and GT35. GT35 pyridoxal phosphate-dependent glycogen phosphorylases share structural and mechanistic similarities with typical NDP-dependent GTs.

== Family Firsts ==
;First sterochemistry determination:
Cellobiose phosphorylase from ''Clostridium thermocellum'' <cite>REF5</cite>
;First gene cloning:
Cellobiose phosphorylase and a cellodextrin phosphorylase from ''Clostridium stercorarium'' <cite>REF6</cite>
;First catalytic nucleophile identification:
The inverting phosphorolytic reaction does not require catalytic general base residue, but inorganic phosphate act as a nucleophile.
;First general acid residue identification:
''Vibrio proteolyticus'' chitobiose phosphorylase by kinetic studies with mutants <cite>REF1</cite>
;First 3-D structure:
''Vibrio proteolyticus'' chitobiose phosphorylase <cite>REF1</cite>.

== References ==
<biblio>
#REF1 pmid=15274915
#REF2 pmid=14756551
#REF3 pmid=11587643
#REF4 pmid=19124470
#REF5 Sih CJ, and McBee RH. ''A cellobiose phosphorylase in Clostridium thermocellum.'' Proc Montana Acad Sci 1955, 15, 21-22.

#REF6 pmid=9249035
#REF7 pmid=17921247

</biblio>

[[Category:Glycoside Hydrolase Families]]

Glycoside Hydrolase Family 94

2009-07-14T14:55:29Z

Masafumi Hidaka: /* Three-dimensional structures */

* [[Author]]: [[User:Masafumi_Hidaka|Masafumi Hidaka]]
* [[Responsible Curator]]: [[User:ShinyaFushinobu|Shinya Fushinobu]]
----

<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Glycoside Hydrolase Family 94'''
|-
|'''Clan'''
|none (similar to GH-L)
|-
|'''Mechanism'''
|inverting
|-
|'''Active site residues'''
|known
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |http://www.cazy.org/fam/GH94.html
|}
</div>

== Substrate specificities ==
This family contains phosphorolytic enzymes (usually named using a combination of “the substrate” and “phosphorylase”) that cleave beta glycosidic bond. The substrate specificities found in GH94 are: cellobiose (Glc-β1,4-Glc) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.20 2.4.1.20]), cellodextrin ((Glc-β1,4-)<sub>n-1</sub>Glc; n ≥ 3) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.29 2.4.1.29]), and N,N’-diacetyl chitobiose (GlcNAc-β1,4-GlcNAc) phosphorylase. Moreover, a phosphorylase domain belonging to this family is found in cyclic β-1,2-glucan synthase (EC 2.4.1.-) along with a [[GlycosylTransferase Family 84]] domain <cite>REF7</cite>. This domain is thought to phosphorolyze protein-bound β-1,2-glucans.<BR> '''Phosphorylases''' catalyze the phosphorolysis of glycosidic bonds to generate glycosyl-phosphate. The reaction is reversible due to the energy of the glycosyl-phosphate bond. Therefore, phosphorylases are categorized as “transferase” among enzyme nomenclature (EC 2.4.1.-). Together with the fact that the GH94 enzymes did not show hydrolytic activity, GH94 enzymes were initially classified in [[GlycosylTransferase Family 36]]. However because of the evolutionary, structural and mechanistic relatedness of GH94 phosphorylases with clan GH-L glycoside hydrolases, the family was re-assigned to family GH94 <cite>REF1</cite>.

== Kinetics and Mechanism ==
Phosphorolysis by GH94 enzymes proceeds with inversion of anomeric configuration, as first shown by Sih and McBee <cite>REF5</cite> on cellobiose phosphorylase from ''Clostridium thermocellum'', i.e. cellobiose (Glc-β1,4-Glc) + Pi ↔ α-glucose 1-phosphate + glucose. Considering the topology of the active site structure, the reaction mechanism for inverting phosphorylase is proposed to be similar to that for inverting GH <cite>REF1</cite>. With the aid of general acid residue, the enzymatic phosphorolysis begins with direct nucleophilic attack by phosphate on the anomeric C-1 carbon, instead of the water molecule activated by a general base residue in inverting GH reaction.

== Catalytic Residues ==
The catalytic residue was firstly estimated by superimposing the active site structure of chitobiose phosphorylase from ''Vibrio proteolyticus'' with a [[Glycoside Hydrolase Family 15]] enzyme, glucoamylase from ''Thermoanaerobacterium thermosaccharolyticum'' <cite>REF1</cite>. Considering the similarities of the active site structure, Asp492 was estimated as the general acid residue. D492A/N mutants of this enzyme showed no detectable activity. General base residue is not required for the reaction of glycoside hydrolase-like inverting phosphorylases.

== Three-dimensional structures ==
The first solved 3-D structure was chitobiose phosphorylase from ''Vibrio proteolyticus'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7V 1V7V], [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7W 1V7W],
[http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7X 1V7X]) <cite>REF1</cite>. The enzyme has a (α/α)<sub>6</sub> barrel fold that is remarkably similar to clan GH-L. The position of the catalytic general acid is superimposable with Clan GH-L. It should be noted that GH94 enzymes act on β-bonds, whereas clan GH-L enzymes (GH15 and GH65) act on α-bonds. <BR> Today, phosphorylases are categorized based on the evolutionary origins. GH type phosphorylases are classified in [[Glycoside Hydrolase Family 13]], [[Glycoside Hydrolase Family 65]], GH94, and [[Glycoside Hydrolase Family 112]]. GH13 sucrose phosphorylase from ''Bifidobacterium adolescentis'' has a TIM barrel fold catalytic domain like other GH13 hydorolytic enzymes (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1R7A 1R7A]) <cite>REF2</cite>. GH65 maltose phorphorylase from ''Lactobacillus brevis'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1H54 1H54]) <cite>REF3</cite> and GH94 enzymes share clan GH-L like (α/α)<sub>6</sub> barrel fold domain. GH112 galacto-''N''-biose/lacto-''N''-biose I phosphorylase from ''Bifidobacterium longum'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUS 2ZUS], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUT 2ZUT], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUU 2ZUU], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUV 2ZUV], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUW 2ZUW] ), which catalyzes phosphorolysis of β-galactosidic bond, has a TIM barrel fold domain similar with that of GH42 β-galactosidase, hydrolase for β-galactosidic bond <cite>REF4</cite>. GT-type phosphorylases are classified in GT4 and GT35. GT35 pyridoxal phosphate-dependent glycogen phosphorylases share structural and mechanistic similarities with typical NDP-dependent GTs.

== Family Firsts ==
;First sterochemistry determination:
Cellobiose phosphorylase from ''Clostridium thermocellum'' <cite>REF5</cite>
;First gene cloning:
Cellobiose phosphorylase and a cellodextrin phosphorylase from ''Clostridium stercorarium'' <cite>REF6</cite>
;First catalytic nucleophile identification:
The inverting phosphorolytic reaction does not require catalytic general base residue, but inorganic phosphate act as a nucleophile.
;First general acid residue identification:
''Vibrio proteolyticus'' chitobiose phosphorylase by kinetic studies with mutants <cite>REF1</cite>
;First 3-D structure:
''Vibrio proteolyticus'' chitobiose phosphorylase <cite>REF1</cite>.

== References ==
<biblio>
#REF1 pmid=15274915
#REF2 pmid=14756551
#REF3 pmid=11587643
#REF4 pmid=19124470
#REF5 Sih CJ, and McBee RH. ''A cellobiose phosphorylase in Clostridium thermocellum.'' Proc Montana Acad Sci 1955, 15, 21-22.

#REF6 pmid=9249035
#REF7 pmid=17921247

</biblio>

[[Category:Glycoside Hydrolase Families]]

Glycoside Hydrolase Family 94

2009-07-11T14:16:01Z

Masafumi Hidaka:

* [[Author]]: [[User:Masafumi_Hidaka|Masafumi Hidaka]]
* [[Responsible Curator]]: [[User:ShinyaFushinobu|Shinya Fushinobu]]
----

<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Glycoside Hydrolase Family 94'''
|-
|'''Clan'''
|none (similar to GH-L)
|-
|'''Mechanism'''
|inverting
|-
|'''Active site residues'''
|known
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |http://www.cazy.org/fam/GH94.html
|}
</div>

== Substrate specificities ==
This family contains phosphorolytic enzymes (usually named using a combination of “the substrate” and “phosphorylase”) that cleave beta glycosidic bond. The substrate specificities found in GH94 are: cellobiose (Glc-β1,4-Glc) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.20 2.4.1.20]), cellodextrin ((Glc-β1,4-)<sub>n-1</sub>Glc; n≥3) phosphorylase (EC [http://us.expasy.org/cgi-bin/nicezyme.pl?2.4.1.29 2.4.1.29]), (N.N’-diacetyl)chitobiose (GlcNAc-β1,4-GlcNAc) phosphorylase, and a domain phosphorolyzing protein-bound β-1,2-glucan accompanied by cyclic β1,2-glucan synthase(EC 2.4.1.-) belonging to GT84.<BR> '''Phosphorylases''' catalyze the phosphorolysis of glycosidic bonds to generate glycosyl-phosphate. The reaction is reversible due to the energy of the glycosyl-phosphate bond. Therefore, phosphorylases are categorized as “transferase” among enzyme nomenclature (EC 2.4.1.-). Together with the fact that none of GH94 enzymes showed hydrolytic activity, GH94 enzymes were formally classified in [[GlycosylTransferase Family 36]]. By revealing the evolutionary, structural and mechanistic relationship of GH94 pshophorylases with glycoside hydrolase of clan GH-L, the family is re-assigned to a GH family <cite>REF1</cite>.

== Kinetics and Mechanism ==
Phosphorolysis by GH94 enzymes proceeds with inversion of anomeric configuration, as first shown by Sih and McBee <cite>REF5</cite> on cellobiose phosphorylase from ''Clostridium thermocellum'', i.e. cellobiose (Glc-β1,4-Glc) + Pi ↔ α-glucose 1-phosphate + glucose. Considering the topology of the active site structure, the reaction mechanism for inverting phosphorylase is proposed to be similar to that for inverting GH <cite>REF1</cite>. With the aid of general acid residue, the enzymatic phosphorolysis begins with direct nucleophilic attack by phosphate on the anomeric C-1 carbon, instead of the water molecule activated by a general base residue in inverting GH reaction.

== Catalytic Residues ==
The catalytic residue was firstly estimated by superimposing the active site structure of chitobiose phosphorylase from ''Vibrio proteolyticus'' with a [[Glycoside Hydrolase Family 15]] enzyme, glucoamylase from ''Thermoanaerobacterium thermosaccharolyticum'' <cite>REF1</cite>. Considering the similarities of the active site structure, Asp492 was estimated as the general acid residue. D492A/N mutants of this enzyme showed no detectable activity. General base residue is not required for the reaction of glycoside hydrolase-like inverting phosphorylases.

== Three-dimensional structures ==
The first solved 3-D structure was chitobiose phosphorylase from ''Vibrio proteolyticus'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7V 1V7V], [http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7W 1V7W],
[http://www.rcsb.org/pdb/explore/explore.do?structureId=1V7X 1V7X]) <cite>REF1</cite>. The enzyme has a (α/α)<sub>6</sub> barrel fold that is remarkably similar to clan GH-L. The position of the catalytic general acid is superimposable with Clan GH-L. It should be noted that GH94 enzymes act on β-bonds, whereas clan GH-L enzyme (GH15 and GH65) act on α-bonds. <BR> Today, phosphorylases are categorized based on the evolutionary origins. GH type phosphorylases are classified in [[Glycoside Hydrolase Family 13]], [[Glycoside Hydrolase Family 65]], GH94, and [[Glycoside Hydrolase Family 112]]. GH13 sucrose phosphorylase from ''Bifidobacterium adolescentis'' has a TIM barrel fold catalytic domain like other GH13 hydorolytic enzymes (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1R7A 1R7A]) <cite>REF2</cite>. GH65 maltose phorphorylase from Lactobacillus brevis (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=1H54 1H54]) <cite>REF3</cite> and GH94 enzymes share clan GH-L like (α/α)<sub>6</sub> barrel fold domain. GH112 galacto-''N''-biose/lacto-''N''-biose I phosphorylase from ''Bifidobacterium longum'' (PDB [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUS 2ZUS], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUT 2ZUT], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUU 2ZUU], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUV 2ZUV], [http://www.rcsb.org/pdb/explore/explore.do?structureId=2ZUW 2ZUW], ), which catalyzes phosphorolysis of β-galactosidic bond, has a TIM barrel fold domain similar with that of GH42 β-galactosidase, hydrolase for β-galactosidic bond <cite>REF4</cite>. GT-type phosphorylases are classified in GT4 and GT35. GT35 pyridoxal phosphate-dependent glycogen phosphorylases share structural and mechanistic similarities with typical NDP-dependent GTs.

== Family Firsts ==
;First sterochemistry determination:
Cellobiose phosphorylase from ''Clostridium thermocellum'' <cite>REF5</cite>
;First gene cloning:
Cellobiose phosphorylase and a cellodextrin phosphorylase from ''Clostridium stercorarium'' <cite>REF6</cite>
;First catalytic nucleophile identification:
The inverting phosphorolytic reaction does not require catalytic general base residue, but inorganic phosphate act as a nucleophile.
;First general acid residue identification:
''Vibrio proteolyticus'' chitobiose phosphorylase by kinetic studies with mutants <cite>REF1</cite>
;First 3-D structure:
''Vibrio proteolyticus'' chitobiose phosphorylase <cite>REF1</cite>.

== References ==
<biblio>
#REF1 pmid=15274915
#REF2 pmid=14756551
#REF3 pmid=11587643
#REF4 pmid=19124470
#REF5 Sih CJ, and McBee RH. ''A cellobiose phosphorylase in Clostridium thermocellum.'' Proc Montana Acad Sci 1955, 15, 21-22.

#REF6 pmid=9249035

</biblio>

[[Category:Glycoside Hydrolase Families]]

2009-07-11T00:37:56Z

Masafumi Hidaka: /* Three-dimensional structures */

Glycoside Hydrolase Family 94

2009-07-11T00:31:26Z

Masafumi Hidaka: /* Catalytic Residues */

Glycoside Hydrolase Family 94

2009-07-11T00:25:34Z

Masafumi Hidaka: /* Kinetics and Mechanism */

Glycoside Hydrolase Family 94

2009-07-11T00:20:20Z

Masafumi Hidaka: /* Phosphorylases */

Glycoside Hydrolase Family 94

2009-07-11T00:01:13Z

Masafumi Hidaka:

Glycoside Hydrolase Family 94

2009-07-10T23:59:55Z

Masafumi Hidaka: /* Substrate specificities */