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Difference between revisions of "Carbohydrate Binding Module Family 96"

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* [[Author]]: [[User:Wenwen Tao|Wenwen Tao]]
 
* [[Author]]: [[User:Wenwen Tao|Wenwen Tao]]
 
* [[Responsible Curator]]:  [[User:Yaoguang Chang|Yaoguang Chang]]
 
* [[Responsible Curator]]:  [[User:Yaoguang Chang|Yaoguang Chang]]
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== Ligand specificities ==
 
== Ligand specificities ==
Mention here all major natural ligand specificities that are found within a given family (also plant or mammalian origin). Certain linkages and promiscuity would also be mentioned here if biologically relevant.
+
Members of the CBM96 family are present within various CAZyme families, including [[PL8]], [[PL35]], [[GH16]], and [[GH136]], suggesting that they may possess a wide range of diverse activities. TM6-N4, originating from the marine thermophilic bacterium ''Defluviitalea phaphyphila'', and DmCBM96-1/DmCBM96-2, derived from the bacterium ''Dysgonomonas sp.'' HDW5A isolated from ''Hydrophilus acuminatus'' , are currently the characterized members in the CBM96 family. TM6-N4 is associated with the catalytic module of [[PL39]] alginate lyase, and its specific binding capacity to alginate was confirmed by affinity electrophoresis analysis. Isothermal titration calorimetry analysis revealed that TM6-N4 exhibits no significant preference for mannuronate or guluronate oligosaccharides <cite>Ji2023</cite>. Compared with TM6-N4, DmCBM96-1 exhibits a relatively low sequence identity of approximately 32%, and DmCBM96-2 is even lower of approximately 25% <cite>Altschul1997</cite>. DmCBM96-1 and DmCBM96-2, which are consecutive CBMs linked to [[PL8]]_3 chondroitin sulfate (CS) lyase, have been confirmed to bind to CS-A (chondroitin 4-sulfate), CS-C (chondroitin 6-sulfate), and Aj-fCS (consisting of CS-type backbone and sulfated fucose branches), but not to other polyuronic acids such as hyaluronic acid, dermatan sulfate, heparin, pectin, and alginate. Comparatively, the relative affinity of both DmCBM96 proteins to CS-C are higher than CS-A <cite>Liu2024</cite>.
 
 
''Note: Here is an example of how to insert references in the text, together with the "biblio" section below:'' Please see these references for an essential introduction to the CAZy classification system: <cite>DaviesSinnott2008 Cantarel2009</cite>. CBMs, in particular, have been extensively reviewed <cite>Boraston2004 Hashimoto2006 Shoseyov2006 Guillen2010 Armenta2017</cite>.
 
  
 
== Structural Features ==
 
== Structural Features ==
''Content in this section should include, in paragraph form, a description of:''
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[[File:CBM96_Fig.1.png|thumb|350px|right|'''Figure 1. Crystal structure of TM6-N4 ([{{PDBlink}}7vbo PDB 7vbo]).''' The strand, helix, loop and Ca<Sup>2+</Sup> were colored in cyan, yellow, white and green respectively.]]
* '''Fold:''' Structural fold (beta trefoil, beta sandwich, etc.)
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The crystal structures of TM6-N4 (1.35 Å) and DmCBM96-1 (2.20 Å) reveal that they both exhibit a typical β-sandwich fold, comprising two antiparallel β-sheets consisting of 11 β-strands, but differ in helixes. The binding sites of both DmCBM96-1 and TM6-N4 are located within the loop regions. Site-directed mutagenesis assay revealed that the basic amino acids Arg27, Lys45, Arg53, Arg157, and the aromatic amino acid Tyr51 at the binding site of DmCBM96-1 are crucial for CS binding <cite>Liu2024</cite>. While , the amino acids located at the bottom or wall of the  shallow groove of TM6-N4, including Lys10, Lys22, Lys25, Lys27, Lys31, Arg36, and Tyr159, are essential for alginate binding <cite>Ji2023</cite>. Among these, the crucial amino acids Lys45 and Arg53 in DmCBM96-1 correspond to the key amino acids Lys27 and Arg36 in TM6-N4, respectively, and they are highly conserved within the CBM96 family, suggesting their pivotal roles in the ligand binding process. Structural and mutational analyses revealed that TM6-N4 possess long-extended binding grooves for alginate chains, characteristic of type B CBM fold <cite>Ji2023</cite>.
* '''Type:''' Include here Type A, B, or C and properties
 
* '''Features of ligand binding:''' Describe CBM binding pocket location (Side or apex) important residues for binding (W, Y, F, subsites), interact with reducing end, non-reducing end, planar surface or within polysaccharide chains. Include examples pdb codes. Metal ion dependent. Etc.
 
  
 
== Functionalities ==  
 
== Functionalities ==  
''Content in this section should include, in paragraph form, a description of:''
+
Members of the CBM96 family are annotated by dbCAN to be associated with several CAZyme families <cite>Yin2012</cite>, and their presumed physiological role is to facilitate enzyme-carbohydrate interactions and enhance the local concentration of enzymes near substrates, thereby promoting the degradation of polysaccharides. Based on the polysaccharide-binding specificity of the CBM96 family, it may also have potential applications in the study of cellular structures or the detection of pathogenic bacteria. Furthermore, through fusion expression, CBM96 could potentially be utilized to optimize the properties of other CAZymes <cite>Ji2023</cite>.
* '''Functional role of CBM:''' Describe common functional roles such as targeting, disruptive, anchoring, proximity/position on substrate.
 
* '''Most Common Associated Modules:''' 1. Glycoside Hydrolase Activity; 2. Additional Associated Modules (other CBM, FNIII, cohesin, dockerins, expansins, etc.)
 
* '''Novel Applications:'''  Include here if CBM has been used to modify another enzyme, or if a CBM was used to label plant/mammalian tissues? Etc.
 
  
 
== Family Firsts ==
 
== Family Firsts ==
;First Identified
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;First Identified: The TM6-N4 derived from the alginate lyase Dp0100 of marine bacterium ''Defluviitalea phaphyphila'', which was first reported in 2023, represents the first characterized member of the CBM96 family <cite>Ji2023</cite>.
:Insert archetype here, possibly including ''very brief'' synopsis.
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;First Structural Characterization: Alginate-binding protein TM6-N4 is the first member of the CBM96 family to undergo X-ray crystallography structural characterization ([{{PDBlink}}7vbo PDB 7vbo]) <cite>Ji2023</cite>. Subsequently, CS-binding protein DmCBM96-1 was also characterized ([{{PDBlink}}2pqr PDB 2pqr]) <cite>Liu2024</cite>.
;First Structural Characterization
 
:Insert archetype here, possibly including ''very brief'' synopsis.
 
  
 
== References ==
 
== References ==
 
<biblio>
 
<biblio>
#Cantarel2009 pmid=18838391
+
#Ji2023 pmid=36592931
#DaviesSinnott2008 Davies, G.J. and Sinnott, M.L. (2008) Sorting the diverse: the sequence-based classifications of carbohydrate-active enzymes. ''The Biochemist'', vol. 30, no. 4., pp. 26-32. [https://doi.org/10.1042/BIO03004026 DOI:10.1042/BIO03004026].
+
#Liu2024 pmid=38805590
#Boraston2004 pmid=15214846
+
#Yin2012 pmid=22645317
#Hashimoto2006 pmid=17131061
+
#Altschul1997 pmid=9254694
#Shoseyov2006 pmid=16760304
 
#Guillen2010 pmid=19908036
 
#Armenta2017 pmid=28547780
 
 
</biblio>
 
</biblio>
  

Latest revision as of 22:05, 20 November 2024

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CAZy DB link
https://www.cazy.org/CBM96.html

Ligand specificities

Members of the CBM96 family are present within various CAZyme families, including PL8, PL35, GH16, and GH136, suggesting that they may possess a wide range of diverse activities. TM6-N4, originating from the marine thermophilic bacterium Defluviitalea phaphyphila, and DmCBM96-1/DmCBM96-2, derived from the bacterium Dysgonomonas sp. HDW5A isolated from Hydrophilus acuminatus , are currently the characterized members in the CBM96 family. TM6-N4 is associated with the catalytic module of PL39 alginate lyase, and its specific binding capacity to alginate was confirmed by affinity electrophoresis analysis. Isothermal titration calorimetry analysis revealed that TM6-N4 exhibits no significant preference for mannuronate or guluronate oligosaccharides [1]. Compared with TM6-N4, DmCBM96-1 exhibits a relatively low sequence identity of approximately 32%, and DmCBM96-2 is even lower of approximately 25% [2]. DmCBM96-1 and DmCBM96-2, which are consecutive CBMs linked to PL8_3 chondroitin sulfate (CS) lyase, have been confirmed to bind to CS-A (chondroitin 4-sulfate), CS-C (chondroitin 6-sulfate), and Aj-fCS (consisting of CS-type backbone and sulfated fucose branches), but not to other polyuronic acids such as hyaluronic acid, dermatan sulfate, heparin, pectin, and alginate. Comparatively, the relative affinity of both DmCBM96 proteins to CS-C are higher than CS-A [3].

Structural Features

Figure 1. Crystal structure of TM6-N4 (PDB 7vbo). The strand, helix, loop and Ca2+ were colored in cyan, yellow, white and green respectively.

The crystal structures of TM6-N4 (1.35 Å) and DmCBM96-1 (2.20 Å) reveal that they both exhibit a typical β-sandwich fold, comprising two antiparallel β-sheets consisting of 11 β-strands, but differ in helixes. The binding sites of both DmCBM96-1 and TM6-N4 are located within the loop regions. Site-directed mutagenesis assay revealed that the basic amino acids Arg27, Lys45, Arg53, Arg157, and the aromatic amino acid Tyr51 at the binding site of DmCBM96-1 are crucial for CS binding [3]. While , the amino acids located at the bottom or wall of the shallow groove of TM6-N4, including Lys10, Lys22, Lys25, Lys27, Lys31, Arg36, and Tyr159, are essential for alginate binding [1]. Among these, the crucial amino acids Lys45 and Arg53 in DmCBM96-1 correspond to the key amino acids Lys27 and Arg36 in TM6-N4, respectively, and they are highly conserved within the CBM96 family, suggesting their pivotal roles in the ligand binding process. Structural and mutational analyses revealed that TM6-N4 possess long-extended binding grooves for alginate chains, characteristic of type B CBM fold [1].

Functionalities

Members of the CBM96 family are annotated by dbCAN to be associated with several CAZyme families [4], and their presumed physiological role is to facilitate enzyme-carbohydrate interactions and enhance the local concentration of enzymes near substrates, thereby promoting the degradation of polysaccharides. Based on the polysaccharide-binding specificity of the CBM96 family, it may also have potential applications in the study of cellular structures or the detection of pathogenic bacteria. Furthermore, through fusion expression, CBM96 could potentially be utilized to optimize the properties of other CAZymes [1].

Family Firsts

First Identified
The TM6-N4 derived from the alginate lyase Dp0100 of marine bacterium Defluviitalea phaphyphila, which was first reported in 2023, represents the first characterized member of the CBM96 family [1].
First Structural Characterization
Alginate-binding protein TM6-N4 is the first member of the CBM96 family to undergo X-ray crystallography structural characterization (PDB 7vbo) [1]. Subsequently, CS-binding protein DmCBM96-1 was also characterized (PDB 2pqr) [3].

References

  1. Ji S, Tian X, Li X, and She Q. (2023). Identification and structural analysis of a carbohydrate-binding module specific to alginate, a representative of a new family, CBM96. J Biol Chem. 2023;299(2):102854. DOI:10.1016/j.jbc.2022.102854 | PubMed ID:36592931 [Ji2023]
  2. Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, and Lipman DJ. (1997). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 1997;25(17):3389-402. DOI:10.1093/nar/25.17.3389 | PubMed ID:9254694 [Altschul1997]
  3. Liu G, Mei X, Zhang Y, Chen G, Li J, Tao W, Sun M, Zheng L, Chang Y, and Xue C. (2024). Characterization and Structural Analysis of a Novel Carbohydrate-Binding Module from Family 96 with Chondroitin Sulfate-Specific Binding Capacity. J Agric Food Chem. 2024;72(23):13196-13204. DOI:10.1021/acs.jafc.4c00090 | PubMed ID:38805590 [Liu2024]
  4. Yin Y, Mao X, Yang J, Chen X, Mao F, and Xu Y. (2012). dbCAN: a web resource for automated carbohydrate-active enzyme annotation. Nucleic Acids Res. 2012;40(Web Server issue):W445-51. DOI:10.1093/nar/gks479 | PubMed ID:22645317 [Yin2012]

All Medline abstracts: PubMed