Glycoside Hydrolase Family 3 - Revision history

Harry Brumer at 23:19, 6 January 2023

2023-01-06T23:19:59Z

Harry Brumer at 23:13, 6 January 2023

2023-01-06T23:13:03Z

Harry Brumer: Finally updated Family First stereochemistry determination

2023-01-06T23:03:18Z

Finally updated Family First stereochemistry determination

Harry Brumer: Text replacement - "\^\^\^(.*)\^\^\^" to "$1"

2021-12-18T21:14:00Z

Text replacement - "\^\^\^(.*)\^\^\^" to "$1"

Harry Brumer: /* Catalytic acid/base */

2017-03-06T20:43:20Z

Catalytic acid/base

Harry Brumer: /* Substrate specificities */

2017-03-06T16:39:45Z

Substrate specificities

Harry Brumer: /* Substrate specificities */

2017-03-06T16:39:11Z

Substrate specificities

Harry Brumer at 16:38, 6 March 2017

2017-03-06T16:38:41Z

Harry Brumer: /* Substrate specificities */

2017-03-06T16:26:44Z

Substrate specificities

Harry Brumer: /* Catalytic acid/base */

2017-03-06T16:23:43Z

Catalytic acid/base

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 == Kinetics and Mechanism ==
 [[Image:GH3_Fig_1.png|thumb|right|350px|'''Figure 1. Ribbon representation of barley β-glucan exohydrolase isoenzyme ExoI.'''  Domain 1, domain 2, and the linker region of the enzyme are coloured in magenta, cyan, and yellow, respectively. Figure from <cite>Harvey2000</cite>.]]
-GH3 [[glycoside hydrolases]] remove single glycosyl residues from the non-reducing ends of their substrates. Catalysis occurs via a [[classical Koshland double-displacement mechanism]] with the anomeric configuration of the released glycose being [[retaining|retained]]. The retention of anomeric configuration has been established experimentally for several enzymes (see <cite>Ducatti2016 Macdonald2015</cite> and references therein). The active site of GH3 enzymes consists of two glucosyl-binding subsites (-1 and +1) with an enzymic nucleophile and general acid/base residue flanking the junction of these two subsites (see <cite>Davies1997</cite> for subsite nomenclature).  This arrangement of catalytic residues was first visualized in 1999 when the crystal structure of barley β-D-glucan glucohydrolase was determined in complex with glucose <cite>Vargese1999</cite> (Fig. 1).
+GH3 [[glycoside hydrolases]] remove single glycosyl residues from the non-reducing ends of their substrates. Catalysis occurs via a [[classical Koshland double-displacement mechanism]] with the anomeric configuration of the released glycose being [[retaining|retained]]. The retention of anomeric configuration has been established experimentally for several enzymes (see <cite>Dan2000 Choengpanya2015 Macdonald2015 Ducatti2016</cite> and references therein). The active site of GH3 enzymes consists of two glucosyl-binding subsites (-1 and +1) with an enzymic nucleophile and general acid/base residue flanking the junction of these two subsites (see <cite>Davies1997</cite> for subsite nomenclature).  This arrangement of catalytic residues was first visualized in 1999 when the crystal structure of barley β-D-glucan glucohydrolase was determined in complex with glucose <cite>Vargese1999</cite> (Fig. 1).
 Several studies have contributed to the understanding of the kinetics and mechanism of GH3 enzymes, including detailed analyses of β-glucosidases from fungi (''Aspergillus wentii ''<cite>Bause1974</cite> and ''Aspergillus niger'' <cite>Dan2000 Thongpoo2013</cite>) and Gram-negative bacteria (''Flavobacterium meningosepticum'' <cite>Chir2002 Li2002</cite>, ''Thermotoga neapolitana'' <cite>Pozzo2010</cite>), as well as a GH3 glucosylceramidase from the Gram-positive microbe ''Paenibacillus ''sp. TS12 <cite>Paal2004</cite>.  Kinetic and mechanistic analyses of β-D-glucan glucohydrolases and two ‘bifunctional’ α-L-arabinofuranosidase/β-D-xylopyranosidases from plants (barley) have also been carried out <cite>Lee2003 Hrmova1998</cite>.  These studies, combined with kinetic and mechanistic analyses of ''N''-acetyl-β-D-glucosaminidases from the Gram-positive microbe ''Bacilus subtillus ''<cite>Litzinger2010b Bacik2012</cite>, and Gram-negative microbes ''Vibrio furnisii'' <cite>Vocadlo2000 Vocadlo2005</cite>, ''Vibrio cholerea'' <cite>Stubbs2007</cite> and ''Salmonella typhimerium'' <cite>Bacik2012</cite> confirm that while the catalytic nucleophile of GH3 enzymes is well conserved, the location and identity of the general acid/base residue is not (see below).
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 #Macdonald2015 pmid=25533455
 #Ducatti2016 pmid=27744113
+#Choengpanya2015 pmid=26166179
 #Legler1979 pmid=389631
 </biblio>
 [[Category:Glycoside Hydrolase Families|GH003]]

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 == Family Firsts ==
-;First stereochemistry determination: Retention of the anomeric configuration during hydrolysis catalyzed by GH3 was first inferred from the early work of Legler on an ''Aspergillus wentii'' beta-glucosidase (see <cite>Dan2000</cite> and references therein). Probably the first direct demonstration by H-1 NMR of retention is the work of Withers, Shoshoyev, et al. on an ''Aspergillus niger'' orthologue <cite>Dan2000</cite>. Retention in a GH3 phosphorylase was first shown for a beta-''N''-acetylglucosaminidase from ''Cellulomonas fimi'' <cite>Macdonald2015</cite>.
+;First stereochemistry determination: Retention of the anomeric configuration during hydrolysis catalyzed by GH3 was first inferred from the early work of Legler et al. on an ''Aspergillus wentii'' beta-glucosidase <cite>Legler1979</cite> (discussed in <cite>Dan2000</cite>). Probably the first direct demonstration by H-1 NMR of retention is the work of Withers, Shoshoyev, et al. on an ''Aspergillus niger'' orthologue <cite>Dan2000</cite>. Retention in a GH3 phosphorylase was first shown for a beta-''N''-acetylglucosaminidase from ''Cellulomonas fimi'' <cite>Macdonald2015</cite>.
 ;First [[catalytic nucleophile]] identification: First suggested by Bause and Legler in 1974 using conduritol B-epoxide labelling of an ''Aspergillus wentii'' glucosidase  <cite>Bause1974</cite>, and later supported by the crystal structures of a product complex of a barley β-D-glucan glucohydrolase <cite>Vargese1999</cite> and a trapped covalent intermediate <cite>Hrmova2001</cite>.  Contemporaneous active-site labeling of an ''A. niger'' β-glucosidase <cite>Dan2000</cite> and ''V. furnisii'' NagZ <cite>Vocadlo2000</cite> using 2-deoxy-2-fluoro-β-D-glycosides allowed unequivocal identification of the catalytic nucleophiles in these enzymes.
@@ Line 130: / Line 130: @@
 #Macdonald2015 pmid=25533455
 #Ducatti2016 pmid=27744113
 </biblio>
 [[Category:Glycoside Hydrolase Families|GH003]]

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 == Family Firsts ==
-;First sterochemistry determination: ''To be added.''
+;First stereochemistry determination: Retention of the anomeric configuration during hydrolysis catalyzed by GH3 was first inferred from the early work of Legler on an ''Aspergillus wentii'' beta-glucosidase (see <cite>Dan2000</cite> and references therein). Probably the first direct demonstration by H-1 NMR of retention is the work of Withers, Shoshoyev, et al. on an ''Aspergillus niger'' orthologue <cite>Dan2000</cite>. Retention in a GH3 phosphorylase was first shown for a beta-''N''-acetylglucosaminidase from ''Cellulomonas fimi'' <cite>Macdonald2015</cite>.
 ;First [[catalytic nucleophile]] identification: First suggested by Bause and Legler in 1974 using conduritol B-epoxide labelling of an ''Aspergillus wentii'' glucosidase  <cite>Bause1974</cite>, and later supported by the crystal structures of a product complex of a barley β-D-glucan glucohydrolase <cite>Vargese1999</cite> and a trapped covalent intermediate <cite>Hrmova2001</cite>.  Contemporaneous active-site labeling of an ''A. niger'' β-glucosidase <cite>Dan2000</cite> and ''V. furnisii'' NagZ <cite>Vocadlo2000</cite> using 2-deoxy-2-fluoro-β-D-glycosides allowed unequivocal identification of the catalytic nucleophiles in these enzymes.

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 {{CuratorApproved}}
-* [[Author]]s: ^^^Geoff Fincher^^^, ^^^Brian Mark^^^, and ^^^Harry Brumer^^^
+* [[Author]]s: [[User:Geoff Fincher|Geoff Fincher]], [[User:Brian Mark|Brian Mark]], and [[User:Harry Brumer|Harry Brumer]]
-* [[Responsible Curator]]:  ^^^Bernard Henrissat^^^
+* [[Responsible Curator]]:  [[User:Bernard Henrissat|Bernard Henrissat]]
 ----

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 Later structural studies of bacterial and fungal GH3 enzymes have since shown that while insertion of additional domains is possible (which shifts the location of the glutamic acid general acid-base within the primary sequence), the contribution of two-domains to the active site architecture, as first observed for the barley enzyme, appears to be a core feature of multidomain GH3 β-glucanases <cite>Pozzo2010 Nakatani2012 Yoshida2010</cite>. Superposition of available GH3 β-D-glucan glucohydrolase structures clearly reveals the conservation of this architecture, and directly highlights active-site residue homology in the absence of protein sequence similarity - especially so in the case of the catalytic acid-base <cite>Thongpoo2013</cite> (Fig. 2).
-Notably, GH3 NagZ enzymes represent a significant departure from the above two-domain active site architecture paradigm. A crystal structure of NagZ from ''B. subtilis'', together with kinetic analysis, provided evidence that the catalytic acid-base is an unusual histidine/aspartate dyad that resides within a flexible loop on the catalytic (β/α)<sub>8</sub> barrel, not on a separate domain <cite>Litzinger2010a</cite>. Though the enzyme adopts a two-domain fold similar to the barely β-D-glucan glucohydrolase, the C-terminal domain does not participate in catalysis (Fig. 3).  In fact, most NagZ enzymes from Gram-negative bacteria are single domain enzymes comprised solely of a catalytic (β/α)<sub>8</sub> barrel that contains the conserved aspartate nucleophile and a catalytic histidine/aspartate dyad on flexible loop, as seen for ''B. subtilis ''NagZ <cite>Bacik2012</cite> (Fig. 4).  The flexible loop containing the His/Asp dyad is unique to GH3 NagZ enzymes and can be identified by the consensus motif [KH(F/I)PG(H/L)GXXXX'''D'''(S/T)'''H'''] (catalytic dyad highlighted in boldface) <cite>Vocadlo2000</cite>.  In light of these studies, the residue identification in a ''Clostridium paraputrificum ''N''-acetyl-β-D-glucosaminidase'' (Nag3A) would appear to be unreliable <cite>Li2006</cite>.
+Notably, GH3 NagZ enzymes represent a significant departure from the above two-domain active site architecture paradigm. A crystal structure of NagZ from ''B. subtilis'', together with kinetic analysis, provided evidence that the catalytic acid-base is an unusual histidine/aspartate dyad that resides within a flexible loop on the catalytic (β/α)<sub>8</sub> barrel, not on a separate domain <cite>Litzinger2010a</cite>. Though the enzyme adopts a two-domain fold similar to the barely β-D-glucan glucohydrolase, the C-terminal domain does not participate in catalysis (Fig. 3).  In fact, most NagZ enzymes from Gram-negative bacteria are single domain enzymes comprised solely of a catalytic (β/α)<sub>8</sub> barrel that contains the conserved aspartate nucleophile and a catalytic histidine/aspartate dyad on flexible loop, as seen for ''B. subtilis ''NagZ <cite>Bacik2012</cite> (Fig. 4).  The flexible loop containing the His/Asp dyad is unique to GH3 NagZ enzymes and can be identified by the consensus motif [KH(F/I)PG(H/L)GXXXX'''D'''(S/T)'''H'''] (catalytic dyad highlighted in boldface) <cite>Vocadlo2000</cite>.  In light of these studies, the residue identification in a ''Clostridium paraputrificum'' ''N''-acetyl-β-D-glucosaminidase (Nag3A) would appear to be unreliable <cite>Li2006</cite>.
-In 2015, a study revisiting the mechanism of ''Cellulomonas fimi'' ''N''-acetylglucosaminidase Nag3, which belongs to the same subfamily as the aforementioned enzymes and likewise has the conserved His/Asp dyad, indicated that Nag3 is predominantly a [[Phosphorylases|glycoside phosphorylase]], rather than a [[glycoside hydrolases|glycoside hydrolase]] <cite>Macdonald2015</cite>.  From this study, it was proposed that all members of the NagZ subfamily are phosphorylases, and that the catalytic histidine is employed to avoid Coulombic repulsion with the incoming acceptor substrate, ''viz.'' phosphate <cite>Macdonald2015</cite>.  However, a more recent study has indicated that not all members of this subfamily are [[phosphorylases]], which casts doubt on the generality of this proposal:  The presence of phosphate does not alter the kinetics of a ''Herbaspirillum seropedicae'' SmR1 beta-''N''-acetylglucosaminidase, and only the [[glycoside hydrolases|hydrolysis product]] is observed <cite>Ducatti2016</cite>.
+In 2015, a study revisiting the mechanism of ''Cellulomonas fimi'' ''N''-acetylglucosaminidase Nag3, which belongs to the same subfamily as the aforementioned enzymes and likewise has the conserved His/Asp dyad, indicated that Nag3 is predominantly a [[Phosphorylases|glycoside phosphorylase]], rather than a [[glycoside hydrolases|glycoside hydrolase]] <cite>Macdonald2015</cite>.  From this study, it was proposed that all members of the NagZ subfamily are phosphorylases, and that the catalytic histidine is employed to avoid Coulombic repulsion with the incoming acceptor substrate, ''viz.'' phosphate <cite>Macdonald2015</cite>.  However, a more recent study has indicated that not all members of this subfamily are [[phosphorylases]], which casts doubt on the generality of this proposal:  The presence of phosphate does not alter the kinetics of a ''Herbaspirillum seropedicae'' SmR1 ''N''-acetyl-β-D-glucosaminidase, and only the [[glycoside hydrolases|hydrolysis product]] is observed <cite>Ducatti2016</cite>.
 == Three-dimensional structures ==

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 == Substrate specificities ==
-GH3 currently groups together exo-acting β-D-glucosidases, α-L-arabinofuranosidases, β-D-xylopyranosidases, ''N''-acetyl-β-D-glucosaminidases ([[glycoside hydrolases]]), and ''N''-acetyl-β-D-glucosaminide [[phosphorylases]] <cite>Harvey2000 Macdonald2015</cite>. Widely distributed in bacteria, fungi and plants, GH3 enzymes carry out a range of functions including cellulosic biomass degradation, plant and bacterial cell wall remodeling, energy metabolism and pathogen defense.  In many cases the enzymes have dual or broad substrate specificities with respect to monosaccharide residues, linkage position and chain length of the substrate. For example, there are several well-characterized ‘bifunctional’ enzymes in the family that have both α-L-arabinofuranosidase and β-D-xylopyranosidase activity <cite>Lee2003</cite>, and one characterized example of an ''N''-acetyl-β-D-glucosaminide/β-glucoside [[glycoside hydrolase|hydrolase]]/[[phosphorylases|phosphorylase]] from ''Cellulomonas fimi'' (Nag3) <cite>Mayer2006 Macdonald2015</cite>.  GH3 β-D-glucosidases from barley, which are more precisely referred to as β-D-glucan glucohydrolases, are also broad specificity exo-hydrolases that remove single glucosyl residues from the non-reducing ends of a range of β-D-glucans, β-D-oligoglucosides and aryl β-D-glucosides, including (1,3)-β-D-glucans, (1,4)-β-D-glucans, (1,3;1,4)-β-D-glucans and (1,6)-β-D-glucans, 4-nitrophenyl β-D-glucoside, certain cyanogenic β-D-glucosides and some β-D-oligoxyloglucosides <cite>Hrmova1998</cite>.
+Glycoside Hydrolase Family 3 currently groups together exo-acting β-D-glucosidases, α-L-arabinofuranosidases, β-D-xylopyranosidases, ''N''-acetyl-β-D-glucosaminidases ([[glycoside hydrolases]]), and ''N''-acetyl-β-D-glucosaminide [[phosphorylases]] <cite>Harvey2000 Macdonald2015</cite>. Widely distributed in bacteria, fungi and plants, GH3 enzymes carry out a range of functions including cellulosic biomass degradation, plant and bacterial cell wall remodeling, energy metabolism and pathogen defense.  In many cases the enzymes have dual or broad substrate specificities with respect to monosaccharide residues, linkage position and chain length of the substrate. For example, there are several well-characterized ‘bifunctional’ enzymes in the family that have both α-L-arabinofuranosidase and β-D-xylopyranosidase activity <cite>Lee2003</cite>, and one characterized example of an ''N''-acetyl-β-D-glucosaminide/β-glucoside [[glycoside hydrolase|hydrolase]]/[[phosphorylases|phosphorylase]] from ''Cellulomonas fimi'' (Nag3) <cite>Mayer2006 Macdonald2015</cite>.  GH3 β-D-glucosidases from barley, which are more precisely referred to as β-D-glucan glucohydrolases, are also broad specificity exo-hydrolases that remove single glucosyl residues from the non-reducing ends of a range of β-D-glucans, β-D-oligoglucosides and aryl β-D-glucosides, including (1,3)-β-D-glucans, (1,4)-β-D-glucans, (1,3;1,4)-β-D-glucans and (1,6)-β-D-glucans, 4-nitrophenyl β-D-glucoside, certain cyanogenic β-D-glucosides and some β-D-oligoxyloglucosides <cite>Hrmova1998</cite>.
 In contrast to the broad substrate specificities observed for the GH3 enzymes described above, GH3 ''N''-acetyl-β-D-glucosaminidases are selective for ''N''-acetyl-β-D-glucosamine (GlcNAc) <cite>Chitlaru1996 Ducatti2016</cite> (though exceptions exist, e.g. ''Cellulomonas fimi'' Nag3 <cite>Mayer2006</cite>).  A notable GH3 ''N''-acetyl-β-D-glucosaminidase of prokaryotes is NagZ, which participates in bacterial cell wall recycling (for review see <cite>Johnson2013</cite>) by removing GlcNAc from 1,6-anhydroMurNAc-peptides in Gram-negative bacteria <cite>Cheng2000</cite>, or GlcNAc from GlcNAc-MurNAc-peptides in Gram-positive bacteria <cite>Litzinger2010b</cite>.  The NagZ product 1,6-anhydroMurNAc-peptide is an important activator of AmpC β-lactamase hyper-production in several Gram-negative pathogens, making the enzyme of interest as a potential therapeutic target <cite>Mark2011</cite>.

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 The GH3 [[glycoside hydrolase]] family currently groups together exo-acting β-D-glucosidases, α-L-arabinofuranosidases, β-D-xylopyranosidases, ''N''-acetyl-β-D-glucosaminidases, and ''N''-acetyl-β-D-glucosaminide phosphorylases <cite>Harvey2000 Macdonald2015</cite>. Widely distributed in bacteria, fungi and plants, GH3 enzymes carry out a range of functions including cellulosic biomass degradation, plant and bacterial cell wall remodeling, energy metabolism and pathogen defense.  In many cases the enzymes have dual or broad substrate specificities with respect to monosaccharide residues, linkage position and chain length of the substrate. For example, there are several well-characterized ‘bifunctional’ enzymes in the family that have both α-L-arabinofuranosidase and β-D-xylopyranosidase activity <cite>Lee2003</cite>, and one characterized example of an ''N''-acetyl-β-D-glucosaminide/β-glucoside [[glycoside hydrolase|hydrolase]]/[[phosphorylases|phosphorylase]] from ''Cellulomonas fimi'' (Nag3) <cite>Mayer2006 Macdonald2015</cite>.  GH3 β-D-glucosidases from barley, which are more precisely referred to as β-D-glucan glucohydrolases, are also broad specificity exo-hydrolases that remove single glucosyl residues from the non-reducing ends of a range of β-D-glucans, β-D-oligoglucosides and aryl β-D-glucosides, including (1,3)-β-D-glucans, (1,4)-β-D-glucans, (1,3;1,4)-β-D-glucans and (1,6)-β-D-glucans, 4-nitrophenyl β-D-glucoside, certain cyanogenic β-D-glucosides and some β-D-oligoxyloglucosides <cite>Hrmova1998</cite>.
-In contrast to the broad substrate specificities observed for the GH3 enzymes described above, GH3 ''N''-acetyl-β-D-glucosaminidases are selective for ''N''-acetyl-β-D-glucosamine (GlcNAc) <cite>Chitlaru1996</cite> (though exceptions exist, e.g. ''Cellulomonas fimi'' Nag3 <cite>Mayer2006</cite>).  A notable GH3 ''N''-acetyl-β-D-glucosaminidase of prokaryotes is NagZ, which participates in bacterial cell wall recycling (for review see <cite>Johnson2013</cite>) by removing GlcNAc from 1,6-anhydroMurNAc-peptides in Gram-negative bacteria <cite>Cheng2000</cite>, or GlcNAc from GlcNAc-MurNAc-peptides in Gram-positive bacteria <cite>Litzinger2010b</cite>.  The NagZ product 1,6-anhydroMurNAc-peptide is an important activator of AmpC β-lactamase hyper-production in several Gram-negative pathogens, making the enzyme of interest as a potential therapeutic target <cite>Mark2011</cite>.
+In contrast to the broad substrate specificities observed for the GH3 enzymes described above, GH3 ''N''-acetyl-β-D-glucosaminidases are selective for ''N''-acetyl-β-D-glucosamine (GlcNAc) <cite>Chitlaru1996 Ducatti2016</cite> (though exceptions exist, e.g. ''Cellulomonas fimi'' Nag3 <cite>Mayer2006</cite>).  A notable GH3 ''N''-acetyl-β-D-glucosaminidase of prokaryotes is NagZ, which participates in bacterial cell wall recycling (for review see <cite>Johnson2013</cite>) by removing GlcNAc from 1,6-anhydroMurNAc-peptides in Gram-negative bacteria <cite>Cheng2000</cite>, or GlcNAc from GlcNAc-MurNAc-peptides in Gram-positive bacteria <cite>Litzinger2010b</cite>.  The NagZ product 1,6-anhydroMurNAc-peptide is an important activator of AmpC β-lactamase hyper-production in several Gram-negative pathogens, making the enzyme of interest as a potential therapeutic target <cite>Mark2011</cite>.
 Due to the high diversity of protein structural arrangements found among GH3 members (see below), the phylogeny of this family is complex.  Classification of GH3 members into subfamilies has been performed previously <cite>Harvey2000 Cournoyer2003</cite>, however a robust subfamily classification (on par with those for [[GH13]] <cite>Stam2006</cite>, [[GH5]] <cite>Aspeborg2012</cite>, and the polysaccharide lyases <cite>Lombard2010</cite>), is currently not available.  However, as more plant genome sequences are published, it is becoming clear that the GH3 glycoside hydrolases in plants are encoded by multi-gene families.  In both monocot and dicot species, there are 15-18 genes for the GH3 enzymes, while in lower plants such as the Physcomitrella and Selaginella mosses 6-7 genes encoding GH3 glycoside hydrolases have been identified (<cite>Henrissat2001 GeislerLee2006</cite>, see also [http://www.phytozome.org/ Phytozome] <cite>Goodstein2012</cite>). Plant β-''N''-acetylglucosaminidases have not been identified in GH3 thus far.