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Difference between revisions of "Glycoside Hydrolase Family 106"
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| − | * [[Author]]: | + | * [[Author]]: [[User:Ana Luis|Ana Luis]] |
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== Substrate specificities == | == Substrate specificities == | ||
| − | The glycoside hydrolases of this | + | The [[glycoside hydrolases]] of this family are α-L-rhamnosidases ([{{EClink}}3.2.1.40 EC 3.2.1.40]), and is comprised exclusively of bacterial members. The first family GH106 enzyme characterized was Rham from ''Sphingomonas paucimobilis'' FP2001. This enzyme showed activity against p-nitrophenyl α-L-rhamnopyranoside <cite>Miyata2005</cite>. More recently, two ''Bacteroides thetaiotaomicron'' enzymes (BT0986 and BT4145) have been characterized. These enzymes are exo-acting targeting linkages present in pectin polysaccharides. BT0986 cleaves the L-Rha-α-1,2-L-Arap linkage in the terminal region of Ccain B of rhamnogalacturonan II <cite>Ndeh2017</cite>. The enzyme BT4145 targets the L-Rha-α-1,4-D-GalA linkage in the backbone of rhamnogalacturonan I <cite>Luis2018</cite>. |
== Kinetics and Mechanism == | == Kinetics and Mechanism == | ||
| − | NMR experiments showed that GH106 members act by inverting the anomeric configuration of the glycone sugar participating in the scissle glycosidic linkage (inverting mechanism), and thus mediate bond cleave through a single displacement mechanism <cite>Luis2018</cite>. Additionally, GH106 enzymes are Ca<sup>2+</sup> dependent <cite>Ndeh2017</cite>. | + | <sup>1<</sup>NMR experiments showed that GH106 members act by inverting the anomeric configuration of the glycone sugar participating in the scissle glycosidic linkage (inverting mechanism), and thus mediate bond cleave through a single displacement mechanism <cite>Luis2018</cite>. Additionally, GH106 enzymes are Ca<sup>2+</sup> dependent <cite>Ndeh2017</cite>. Other ion dependent enzyme families are the exo-α-mannosidases [[GH38]], [[GH47]] and [[GH92]]. The structure of BT0986 in complex with D-rhamnopyranose-tetrazole indicates that catalysis is proceeds via a ''B''<sub>2,5</sub> transition state <cite>Ndeh2017</cite>. |
== Catalytic Residues == | == Catalytic Residues == | ||
[[File:BT0986_2.png|thumb|300px|right|'''Figure 1.''' '''BT0986 active site.''' The catalytic amino acids are shown in orange and yellow are the remaining key residues present in the active site. The calcium ion in the active site is shown as a cyan sphere and its polar contacts indicated by black dashed lines. D-rhamnopyranose tetrazole is represented in green.]] | [[File:BT0986_2.png|thumb|300px|right|'''Figure 1.''' '''BT0986 active site.''' The catalytic amino acids are shown in orange and yellow are the remaining key residues present in the active site. The calcium ion in the active site is shown as a cyan sphere and its polar contacts indicated by black dashed lines. D-rhamnopyranose tetrazole is represented in green.]] | ||
| − | Structural characterization of ''B. thetaiotaomicron'' BT0986 identified two carboxylate amino acids (Glu461 and Glu593) that were essential for activity and are highly conserved within GH106 (Figure 1). These glutamates are separated by 8.0 Å, a distance between catalytic residues consistent with an inverting mechanism. Additionally, Glu593, which is 5.8 Å from the anomeric carbon, is ideally positioned to act as general base and Glu461 is the general acid <cite>Ndeh2017</cite>. | + | Structural characterization of ''B. thetaiotaomicron'' BT0986 identified two carboxylate amino acids (Glu461 and Glu593) that were essential for activity and are highly conserved within GH106 (Figure 1). These glutamates are separated by 8.0 Å, a distance between catalytic residues consistent with an inverting mechanism. Additionally, Glu593, which is 5.8 Å from the anomeric carbon, is ideally positioned to act as general base and Glu461 is the likely general acid as it is within hydrogen bonding distance with the glycosidic oxygen <cite>Ndeh2017</cite>. |
== Three-dimensional structures == | == Three-dimensional structures == | ||
| − | [[File:BT0986_1.png|thumb|300px|right|'''Figure 2.''' '''BT0986 structure'''([ | + | [[File:BT0986_1.png|thumb|300px|right|'''Figure 2.''' '''BT0986 structure'''([{{PDBlink}}5MQN 5MQN]). The figure shows the (β/α)<sub>8</sub>-barrel domain (1, yellow) and the additional two β-stranded domains (2 to 6). The D-rhamnopyranose tetrazole represented as sticks with carbon in green and the calcium ion (cyan sphere) are shown at the centre of the (β/α)<sub>8</sub>-barrel.]] |
| − | + | The three-dimensional structure of ''B. thetaiotaomicron'' BT0986 solved using X-ray crystallography represents the first structure of an GH106 enzyme ([{{PDBlink}}5MQN 5MQN]). BT0986 displays a N-terminal catalytic domain that presents an (β/α)<sub>8</sub>-barrel fold with several appended β-stranded domains (C-terminal) (Figure 2) <cite>Ndeh2017</cite>. Calcium makes polar interactions with O2 and O3 of rhamnose (Figure 1) and plays a role in substrate binding and the stabilization of the transition state. | |
== Family Firsts == | == Family Firsts == | ||
Latest revision as of 13:15, 18 December 2021
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| Glycoside Hydrolase Family GH106 | |
| Clan | none |
| Mechanism | inverting |
| Active site residues | known |
| CAZy DB link | |
| https://www.cazy.org/GH106.html | |
Substrate specificities
The glycoside hydrolases of this family are α-L-rhamnosidases (EC 3.2.1.40), and is comprised exclusively of bacterial members. The first family GH106 enzyme characterized was Rham from Sphingomonas paucimobilis FP2001. This enzyme showed activity against p-nitrophenyl α-L-rhamnopyranoside [1]. More recently, two Bacteroides thetaiotaomicron enzymes (BT0986 and BT4145) have been characterized. These enzymes are exo-acting targeting linkages present in pectin polysaccharides. BT0986 cleaves the L-Rha-α-1,2-L-Arap linkage in the terminal region of Ccain B of rhamnogalacturonan II [2]. The enzyme BT4145 targets the L-Rha-α-1,4-D-GalA linkage in the backbone of rhamnogalacturonan I [3].
Kinetics and Mechanism
1<NMR experiments showed that GH106 members act by inverting the anomeric configuration of the glycone sugar participating in the scissle glycosidic linkage (inverting mechanism), and thus mediate bond cleave through a single displacement mechanism [3]. Additionally, GH106 enzymes are Ca2+ dependent [2]. Other ion dependent enzyme families are the exo-α-mannosidases GH38, GH47 and GH92. The structure of BT0986 in complex with D-rhamnopyranose-tetrazole indicates that catalysis is proceeds via a B2,5 transition state [2].
Catalytic Residues
Structural characterization of B. thetaiotaomicron BT0986 identified two carboxylate amino acids (Glu461 and Glu593) that were essential for activity and are highly conserved within GH106 (Figure 1). These glutamates are separated by 8.0 Å, a distance between catalytic residues consistent with an inverting mechanism. Additionally, Glu593, which is 5.8 Å from the anomeric carbon, is ideally positioned to act as general base and Glu461 is the likely general acid as it is within hydrogen bonding distance with the glycosidic oxygen [2].
Three-dimensional structures
The three-dimensional structure of B. thetaiotaomicron BT0986 solved using X-ray crystallography represents the first structure of an GH106 enzyme (5MQN). BT0986 displays a N-terminal catalytic domain that presents an (β/α)8-barrel fold with several appended β-stranded domains (C-terminal) (Figure 2) [2]. Calcium makes polar interactions with O2 and O3 of rhamnose (Figure 1) and plays a role in substrate binding and the stabilization of the transition state.
Family Firsts
- First stereochemistry determination
- BT4145 from Bacteroides thetaiotaomicron [3].
- First catalytic nucleophile identification
- BT0986 from Bacteroides thetaiotaomicron [2].
- First general acid/base residue identification
- BT0986 from Bacteroides thetaiotaomicron [2].
- First 3-D structure
- BT0986 from Bacteroides thetaiotaomicron [2].
References
-
Miyata T, Kashige N, Satho T, Yamaguchi T, Aso Y and Miake F.Cloning (2005) Sequence analysis, and expression of the gene encoding Sphingomonas paucimobilis FP2001 alpha-L-rhamnosidase. Curr Microbiol, vol 51, no. 2., pp. 105-109.
- Ndeh D, Rogowski A, Cartmell A, Luis AS, Baslé A, Gray J, Venditto I, Briggs J, Zhang X, Labourel A, Terrapon N, Buffetto F, Nepogodiev S, Xiao Y, Field RA, Zhu Y, O'Neil MA, Urbanowicz BR, York WS, Davies GJ, Abbott DW, Ralet MC, Martens EC, Henrissat B, and Gilbert HJ. (2017). Complex pectin metabolism by gut bacteria reveals novel catalytic functions. Nature. 2017;544(7648):65-70. DOI:10.1038/nature21725 |
- Luis AS, Briggs J, Zhang X, Farnell B, Ndeh D, Labourel A, Baslé A, Cartmell A, Terrapon N, Stott K, Lowe EC, McLean R, Shearer K, Schückel J, Venditto I, Ralet MC, Henrissat B, Martens EC, Mosimann SC, Abbott DW, and Gilbert HJ. (2018). Dietary pectic glycans are degraded by coordinated enzyme pathways in human colonic Bacteroides. Nat Microbiol. 2018;3(2):210-219. DOI:10.1038/s41564-017-0079-1 |