CAZypedia - User contributions [en-ca]

User:Stefen Stangherlin

2020-08-28T13:54:50Z

Stefen Stangherlin:

[[Image:Stefen.jpg|200px|right]]
Stefen Stangherlin received his BScH in Chemistry from Wilfrid Laurier University in 2019, where he studied the synthesis of plasmonic metal nanoparticles in the lab of Dr. Vladimir Kitaev <cite>Stangherlin2020</cite>. Currently, Stefen is an MSc candidate in the lab of Dr. ^^^Anthony J. Clarke^^^ at Wilfrid Laurier University, where he studies the discovery and development of inhibitors of peptidoglycan ''O''-acetyltransferase B (PatB) in ''Neisseria gonnorhoeae''.

----

<biblio>
Stangherlin2020 Stangherlin, S.; Cathcart, N.; Sato, F.; Kitaev, V. Gold Nanoprisms: Synthetic Approaches for Mastering Plasmonic Properties and Implications for Biomedical Applications. ACS Appl. Nano Mater. 2020, 3 (8), 8304–8318. DOI: 10.1021/acsanm.0c01741
</biblio>


[[Category:Contributors|Stangherlin,Stefen]]

Carbohydrate Esterase Family 3

2020-06-16T22:50:30Z

Stefen Stangherlin: /* Kinetics and Mechanism */

{{CuratorApproved}}
* [[Author]]: ^^^Stefen Stangherlin^^^
* [[Responsible Curator]]s: ^^^Michael Suits^^^ and ^^^Joel Weadge^^^
----


<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Carbohydrate Esterase Family CE3'''
|-
|'''Clan'''
|(α/β/α)-Sandwich
|-
|'''Mechanism'''

|Serine Hydrolase

|-
|'''Active site residues'''
|Known, Catalytic Triad
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |{{CAZyDBlink}}CE3.html
|}
</div>


== Substrate specificities ==
Carbohydrate Esterase Family 3 is currently comprised entirely of de-''O''-acetylxylan esterases. Xylan is a plant cell-wall polysaccharide composed of β-1,4-linked xylose decorated with α-arabinofuranose and α-glucuronic acid substituents <cite>Faik2010</cite>.
== Catalytic Residues ==
Functionally characterized CE3 members are all known to contain the classical catalytic triad of Ser-His-Asp, typical of the SGNH hydrolase family of enzymes (see Fig. 1) <cite>Polgar2005 Molgaard2000</cite>. The active site residues are presented via four conserved consensus sequences (Blocks I-III and V), and have an altered nucleophilic “elbow” turn motif (-GxSxT- as opposed to the canonical -GxSxG- motif) compared to other related members of the α/β-hydrolase family <cite>UptonBuckley1995 Akoh2004</cite>. The catalytic triad along with the Block II Gly and Block III Asn residues that comprise the oxyanion hole, are universally conserved across all characterized CE3 enzymes. The Block V Asp residue mediates the amphoteric nature of the Block V His residue, which abstracts a proton from the Block I Ser to render it nucleophilic.[[Image:CE3_Figure.png|thumb|300px|Figure 1:''Tc''AE206 from ''Talaromyces cellulolyticus'' ([{{PDBlink}}5B5S PDB ID 5B5S]). Colours correspond to α-helices (cyan), β-sheets (magenta), loops (wheat), disulfide bond (yellow), calcium ion (orange) and the active site residues (green).]]

== Kinetics and Mechanism ==
CE3 esterases catalyze the hydrolysis of O-linked acetyl groups from xylan oligo- and poly-saccharides. The Block V His residue abstracts a proton from the Block I Ser, rendering it nucleophilic, which attacks the electrophilic carbonyl carbon of the acetyl group of the xylan substrate; generating a tetrahedral oxyanion intermediate that is stabilized by the backbone amides of the Block I Ser and Block II Gly, as well as the sidechain amide of the Block III Asn, together forming the oxyanion hole in the active site <cite>Uechi2016</cite>. Collapse of the oxyanion intermediate results in the formation of a transient acyl-enzyme intermediate and alcohol by-product <cite>Uechi2016</cite>. A hydrolytic water molecule is then deprotonated by the Block V His residue, and attacks the acyl-enzyme intermediate; hydrolyzing the bond and releasing acetate and the free enzyme <cite>Uechi2016</cite>. In the process, the Ser (Block I) is re-protonated and ready for another catalytic cycle.

The kinetics for two enzymes from the CE3 family have been studied. ''Ct''Ces3-1 ([{{PDBlink}}2VPT PDB ID 2VPT]) was found to have a ''k''cat/''K''M of 2.5 and 1.2 mM-1s-1 for ''p''-nitrophenyl acetate (''p''NP-Ac) and acetylated xylan, respectively <cite>Correia2008</cite>. In other studies, ''Tc''AE206 ([{{PDBlink}}5B5S PDB ID 5B5S]) was not assayed against acetylated xylan, but the ''k''cat/''K''M with ''p''NP-Ac and ''p''-nitrophenyl butyrate (''p''NP-B) were reported as 44.7 and 4.1 mM-1s-1, respectively, while no activity was detected with ''p''‐nitrophenyl octanoate (''p''NP-O) as the substrate <cite>Watanabe2015</cite>.

== Three-dimensional structures ==
Two members of the CE3 family have been structurally resolved, ''Tc''AE206 from ''Talaromyces cellulolyticus'' ([{{PDBlink}}5B5S PDB ID 5B5S]) (see Fig. 1) and ''Ct''Ces3-1 from ''Hungateiclostridium thermocellum'' (formerly ''Clostridium thermocellum'') ([{{PDBlink}}2VPT PDB ID 2VPT]). Both structures adopt an (α/β/α)-sandwich fold typical of the SGNH hydrolase family. The (α/β/α)-sandwich contains five central parallel β-strands forming a curved β-sheet, which is flanked by 5-6 α-helices <cite>Correia2008 Uechi2016</cite>. Additionally, both structures contain a calcium binding loop motif (DXVGX7DXn(D/N)) located above the N-terminal end of the central β-strand (β2) <cite>Watanabe2015</cite>. This binding motif is conserved across all currently characterized CE3s. A coordinated zinc ion was also observed next to a calcium ion in a ''Tc''AE206_S10A variant ([{{PDBlink}}5B5L PDB ID 5B5L]), however this was attributed to the use of ZnSO4 in the crystallization conditions <cite>Uechi2016</cite>. Unique to ''Tc''AE206 is a disulfide bond formed near the N-terminus (see Fig. 1) that is thought to position the catalytic Ser by stabilizing neighbouring areas, including a β-turn (β1) that involves the catalytic Ser <cite>Uechi2016 Watanabe2015</cite>.

== Family Firsts ==
;First characterized: In 1994, the sequence of XynB from ''Ruminococcus flavefaciens'' 17 was found to be related to family G xylanases <cite>Zhang1994</cite>. In 1997, BnaC from ''Neocallimastix patriciarum'' was found to have close relation to XynB and other enzymes known to be members of a diverse family of esterases <cite>Dalrymple1997</cite>. It wasn’t until 2000 that CesA from ''R. flavefaciens'' 17, which was shown to have significant sequence identity to XynB, was characterized with the ability to deacetylate acetylated xylans; thereby representing the first characterized enzymes of family 3 CEs <cite>Aurilia2000</cite>.
;First mechanistic insight: In 2000, CesA, XynB, and BnaC were aligned and shown to contain what was thought to be a Ser-His-Asp catalytic triad responsible for the observed esterase activity <cite>Aurilia2000</cite>. This was later confirmed by the structural resolution of ''Ct''Ces3-1 ([{{PDBlink}}2VPT PDB ID 2VPT]) <cite>Correia2008</cite>.
;First 3-D structure: The first resolved structure was ''Ct''Ces3-1 ([{{PDBlink}}2VPT PDB ID 2VPT]) from ''Hungateiclostridium thermocellum'' (formerly ''Clostridium thermocellum''), displaying the (α/β/α)-sandwich fold and Ser-His-Asp catalytic triad typical of SGNH hydrolases <cite>Correia2008</cite>.

== References ==
<biblio>
#Faik2010 pmid=20375115
#Polgar2005 pmid=16003488
#Molgaard2000 pmid=10801485
#UptonBuckley1995 pmid=7610479
#Akoh2004 pmid=15522763
#Uechi2016 pmid=27329813
#Watanabe2015 pmid=25825334
#Correia2008 pmid=18436237
#Zhang1994 pmid=7816035
#Dalrymple1997 pmid=9274014
#Aurilia2000 pmid=10846217
</biblio>

[[Category:Carbohydrate Esterase Families|CE003]]

Carbohydrate Esterase Family 3

2020-06-16T18:53:10Z

Stefen Stangherlin:

{{CuratorApproved}}
* [[Author]]: ^^^Stefen Stangherlin^^^
* [[Responsible Curator]]s: ^^^Michael Suits^^^ and ^^^Joel Weadge^^^
----


<div style="float:right">
{| {{Prettytable}}
|-
|{{Hl2}} colspan="2" align="center" |'''Carbohydrate Esterase Family CE3'''
|-
|'''Clan'''
|(α/β/α)-Sandwich
|-
|'''Mechanism'''

|Serine Hydrolase

|-
|'''Active site residues'''
|Known, Catalytic Triad
|-
|{{Hl2}} colspan="2" align="center" |'''CAZy DB link'''
|-
| colspan="2" |{{CAZyDBlink}}CE3.html
|}
</div>


== Substrate specificities ==
Carbohydrate Esterase Family 3 is currently comprised entirely of de-''O''-acetylxylan esterases. Xylan is a plant cell-wall polysaccharide composed of β-1,4-linked xylose decorated with α-arabinofuranose and α-glucuronic acid substituents <cite>Faik2010</cite>.
== Catalytic Residues ==
Functionally characterized CE3 members are all known to contain the classical catalytic triad of Ser-His-Asp, typical of the SGNH hydrolase family of enzymes (see Fig. 1) <cite>Polgar2005 Molgaard2000</cite>. The active site residues are presented via four conserved consensus sequences (Blocks I-III and V), and have an altered nucleophilic “elbow” turn motif (-GxSxT- as opposed to the canonical -GxSxG- motif) compared to other related members of the α/β-hydrolase family <cite>UptonBuckley1995 Akoh2004</cite>. The catalytic triad along with the Block II Gly and Block III Asn residues that comprise the oxyanion hole, are universally conserved across all characterized CE3 enzymes. The Block V Asp residue mediates the amphoteric nature of the Block V His residue, which abstracts a proton from the Block I Ser to render it nucleophilic.[[Image:CE3_Figure.png|thumb|300px|Figure 1:''Tc''AE206 from ''Talaromyces cellulolyticus'' ([{{PDBlink}}5B5S PDB ID 5B5S]). Colours correspond to α-helices (cyan), β-sheets (magenta), loops (wheat), disulfide bond (yellow), calcium ion (orange) and the active site residues (green).]]

== Kinetics and Mechanism ==
CE3 esterases catalyze the hydrolysis of O-linked acetyl groups from xylan oligo- and poly-saccharides. The Block V His residue abstracts a proton from the Block I Ser, rendering it nucleophilic, which attacks the electrophilic carbonyl carbon of the acetyl group of the xylan substrate; generating a tetrahedral oxyanion intermediate that is stabilized by the backbone amides of the Block I Ser and Block II Gly, as well as the sidechain amide of the Block III Asn, together forming the oxyanion hole in the active site <cite>Uechi2016</cite>. Collapse of the oxyanion intermediate results in the formation of a transient acyl-enzyme intermediate and alcohol by-product <cite>Uechi2016</cite>. A hydrolytic water molecule is then deprotonated by the Block V His residue, and attacks the acyl-enzyme intermediate; hydrolyzing the bond and releasing acetate and the free enzyme <cite>Uechi2016</cite>. In the process, the Ser (Block I) is re-protonated and ready for another catalytic cycle.

The kinetics for two enzymes from the CE3 family have been studied. ''Ct''Ces3-1 ([{{PDBlink}}2VPT PDB ID 2VPT]) was found to have a ''k''cat/''K''M of 2.5 and 1.2 mM-1s-1 for ''p''-nitrophenyl acetate (''p''NP-Ac) and acetylated xylan, respectively <cite>Correia2008</cite>. In other studies, ''Tc''AE206 ([{{PDBlink}}5B5S PDB ID 5B5S]) was not assayed against acetylated xylan, but the ''k''cat/''K''M with ''p''NP-Ac and ''p''-nitropheyl butyrate (''p''NP-B) were reported as 44.7 and 4.1 mM-1s-1, respectively, while no activity was detected with ''p''‐nitrophenyl octanoate (''p''NP-O) as the substrate <cite>Watanabe2015</cite>.

== Three-dimensional structures ==
Two members of the CE3 family have been structurally resolved, ''Tc''AE206 from ''Talaromyces cellulolyticus'' ([{{PDBlink}}5B5S PDB ID 5B5S]) (see Fig. 1) and ''Ct''Ces3-1 from ''Hungateiclostridium thermocellum'' (formerly ''Clostridium thermocellum'') ([{{PDBlink}}2VPT PDB ID 2VPT]). Both structures adopt an (α/β/α)-sandwich fold typical of the SGNH hydrolase family. The (α/β/α)-sandwich contains five central parallel β-strands forming a curved β-sheet, which is flanked by 5-6 α-helices <cite>Correia2008 Uechi2016</cite>. Additionally, both structures contain a calcium binding loop motif (DXVGX7DXn(D/N)) located above the N-terminal end of the central β-strand (β2) <cite>Watanabe2015</cite>. This binding motif is conserved across all currently characterized CE3s. A coordinated zinc ion was also observed next to a calcium ion in a ''Tc''AE206_S10A variant ([{{PDBlink}}5B5L PDB ID 5B5L]), however this was attributed to the use of ZnSO4 in the crystallization conditions <cite>Uechi2016</cite>. Unique to ''Tc''AE206 is a disulfide bond formed near the N-terminus (see Fig. 1) that is thought to position the catalytic Ser by stabilizing neighbouring areas, including a β-turn (β1) that involves the catalytic Ser <cite>Uechi2016 Watanabe2015</cite>.

== Family Firsts ==
;First characterized: In 1994, the sequence of XynB from ''Ruminococcus flavefaciens'' 17 was found to be related to family G xylanases <cite>Zhang1994</cite>. In 1997, BnaC from ''Neocallimastix patriciarum'' was found to have close relation to XynB and other enzymes known to be members of a diverse family of esterases <cite>Dalrymple1997</cite>. It wasn’t until 2000 that CesA from ''R. flavefaciens'' 17, which was shown to have significant sequence identity to XynB, was characterized with the ability to deacetylate acetylated xylans; thereby representing the first characterized enzymes of family 3 CEs <cite>Aurilia2000</cite>.
;First mechanistic insight: In 2000, CesA, XynB, and BnaC were aligned and shown to contain what was thought to be a Ser-His-Asp catalytic triad responsible for the observed esterase activity <cite>Aurilia2000</cite>. This was later confirmed by the structural resolution of ''Ct''Ces3-1 ([{{PDBlink}}2VPT PDB ID 2VPT]) <cite>Correia2008</cite>.
;First 3-D structure: The first resolved structure was ''Ct''Ces3-1 ([{{PDBlink}}2VPT PDB ID 2VPT]) from ''Hungateiclostridium thermocellum'' (formerly ''Clostridium thermocellum''), displaying the (α/β/α)-sandwich fold and Ser-His-Asp catalytic triad typical of SGNH hydrolases <cite>Correia2008</cite>.

== References ==
<biblio>
#Faik2010 pmid=20375115
#Polgar2005 pmid=16003488
#Molgaard2000 pmid=10801485
#UptonBuckley1995 pmid=7610479
#Akoh2004 pmid=15522763
#Uechi2016 pmid=27329813
#Watanabe2015 pmid=25825334
#Correia2008 pmid=18436237
#Zhang1994 pmid=7816035
#Dalrymple1997 pmid=9274014
#Aurilia2000 pmid=10846217
</biblio>

[[Category:Carbohydrate Esterase Families|CE003]]

User:Stefen Stangherlin

2020-05-28T21:04:16Z

Stefen Stangherlin:

File:Stefen.jpg

2020-05-28T21:02:14Z

Stefen Stangherlin:

User:Stefen Stangherlin

2020-05-28T16:12:31Z

Stefen Stangherlin:

Stefen Stangherlin received his BScH in Chemistry from Wilfrid Laurier University in 2019, where he studied the synthesis of plasmonic metal nanoparticles in the lab of Dr. Vladimir Kitaev. Currently, Stefen is an MSc candidate in the lab of Dr. Anthony J. Clarke at Wilfrid Laurier University, where he studies the discovery and development of inhibitors of peptidoglycan ''O''-acetyltransferase B (PatB) in ''Neisseria gonnorhoeae''.

----


[[Category:Contributors|Stangherlin,Stefen]]

User:Stefen Stangherlin

2020-05-28T16:12:19Z

Stefen Stangherlin:

[[Image:Blank_user-200p.png|200px|right]]
Stefen Stangherlin received his BScH in Chemistry from Wilfrid Laurier University in 2019, where he studied the synthesis of plasmonic metal nanoparticles in the lab of Dr. Vladimir Kitaev. Currently, Stefen is an MSc candidate in the lab of Dr. Anthony J. Clarke at Wilfrid Laurier University, where he studies the discovery and development of inhibitors of peptidoglycan ''O''-acetyltransferase B (PatB) in ''Neisseria gonnorhoeae''.

----


[[Category:Contributors|Stangherlin,Stefen]]

User:Stefen Stangherlin

2020-05-28T15:58:51Z

Stefen Stangherlin:

User:Stefen Stangherlin

2020-05-28T15:58:28Z

Stefen Stangherlin:

User:Stefen Stangherlin

2020-05-28T15:57:10Z

Stefen Stangherlin:

User:Stefen Stangherlin

2020-05-28T15:55:49Z

Stefen Stangherlin:

[[Image:Blank_user-200px.png|200px|right]]
Stefen Stangherlin received his BScH in Chemistry from Wilfrid Laurier University in 2019, where he studied the synthesis of plasmonic metal nanoparticles in the lab of Dr. Vladimir Kitaev. Currently, Stefen is an MSc candidate in the lab of Dr. Anthony J. Clarke at Wilfrid Laurier University, where he studies the discovery and development of inhibitors of peptidoglycan O-acetyltransferase B (PatB) in Neisseria gonnorhoeae.

* See [[User:Gerlind_Sulzenbacher]] for an example. You may copy text from this example by opening the page in another browser window and clicking the "Edit" tab.
* Add your publications in the list below using PubMed IDs and cite them in the text like this <cite>Gilbert2008</cite>.
* Please upload a picture of yourself using the "Upload file" link in the Toolbox section of the left menu, and then replace the Image filename with your own.

''More specific help on these steps is available from the links under the "For contributors" section of the left page menu.''

----

<biblio>
#Gilbert2008 pmid=18430603

</biblio>


[[Category:Contributors|Stangherlin,Stefen]]

User:Stefen Stangherlin

2020-05-27T23:48:42Z

Stefen Stangherlin:

[[Image:Blank_user-200px.png|200px|right]]
'''This is an empty template to help you get started with composing your User page.'''

You should begin by opening this page for editing by clicking on the Edit tab above. Your biography goes in this area of the page.

* See [[User:Gerlind_Sulzenbacher]] for an example. You may copy text from this example by opening the page in another browser window and clicking the "Edit" tab.
* Add your publications in the list below using PubMed IDs and cite them in the text like this <cite>Gilbert2008</cite>.
* Please upload a picture of yourself using the "Upload file" link in the Toolbox section of the left menu, and then replace the Image filename with your own.

''More specific help on these steps is available from the links under the "For contributors" section of the left page menu.''

----

<biblio>
#Gilbert2008 pmid=18430603

</biblio>


[[Category:Contributors|Stangherlin,Stefen]]