Carbohydrate Binding Module Family 41

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• Author: ^^^Alicia Lammerts van Bueren^^^
• Responsible Curator: ^^^Al Boraston^^^

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CAZy DB link

https://www.cazy.org/CBM41.html

Ligand specificities

Modules from family CBM41 bind to alpha-glucans including starch, glycogen, amylose, amylopectin and pullulan, and shorter alpha glucan oligosaccharides derived from these polysaccharides including maltose, maltotriose, longer maltooligosaccharides up to DP7, glucosyl-maltotriose and glucosyl-maltotriosyl-maltotriose [1]. CBM41 modules are specific for alpha-1,4-linked glucose chains but can also accommodate a linear alpha-1,6-linked glucose moiety.

Structural Features

To date there are 11 X-ray crystal structures of CBM41 modules of which seven are in complex with carbohydrate ligand. All adopt a common beta-sandwich fold and form a concave-shaped binding groove on the side of the protein molecule to accommodate the helical structure formed by alpha-1,4-linked maltooligosaccharides. Typically two solvent exposed tryptophan residues form hydrophobic stacking interactions with the primary glucose molecule, with a third tryptophan creating a platform for interacting with longer maltooligosaccharide chains. The binding groove is made up of 4 binding subsites that interact with up to 4 intra-chain alpha-1,4-linked glucose molecules, classifying them as Type B CBMs. The CBM41 module from Thermotoga maritima was shown to accommodate either an alpha-1,4 or alpha-1,6-linked glucose residue in the fourth subsite, demonstrating that there is room for flexibility in the linkage that can be accommodated at this site [2]. The mode of starch binding by CBM41 is similar to other starch-binding CBM families, which include CBM20, 21, , 25, 26, 34, and 48. Although these different starch-binding module families do not share amino-acid sequence similarities with each other, that fact that they share almost identical modes of starch-binding suggests a common evolution towards maltooligosaccharide recognition by all starch-binding CBM families.

There are X-ray crystal structures available for several full-length pullulanases and glycogen-debranching enzymes containing both their catalytic modules and associated CBMs in complex with substrate which has allowed us to understand how modularity of these enzymes contributes to their overall function. Full length glycogen-debranching enzyme SpuA from Streptococcus pneumoniae with its dual tandemly arranged N-terminal CBM41 modules revealed that the most N-terminal CBM41 module directly participates in binding alpha-1,6-linked glucose branch points within the active site of the C-terminal GH13 catalytic module [3]. This is the first demonstration that a CBM directly participates in substrate binding and thus far this feature has only been found within this class of enzyme. The second CBM41 is available to interact with an adjacent alpha-glucan chain, suggesting a possible disruptive role for these CBMs in loosening granular glycogen and increasing the substrate availability for the catalytic module.

Content in this section should include, in paragraph form, a description of:

• Fold: Structural fold (beta trefoil, beta sandwich, etc.)
• 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

Unlike other starch-binding CBM families, CBM41 modules are solely associated with starch debranching enzymes in family GH13, including pullulanases (EC 3.2.1.41 GH13 subfamily 14) and starch/glycogen debranching enzymes (GH13 subfamily 12). The most interesting feature of these modules is they are primarily associated with pullulanase-like enzymes originating from pathogenic bacteria, including pathogenic Streptococcus, Klebsiella and Bacillusspecies. The role of CBM41 is to direct the enzyme onto alpha-1,4-glucan chains and direct the catalytic machinery to alpha-1,6-branch points. Content in this section should include, in paragraph form, a description of:

• 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

First Identified

Family 41 CBMs were previously known as X28 modules. They were first classified as a CBM in 2004 after demonstrating alpha-glucan binding by an N-terminal X28 module from Thermotoga maritima pullulanase PulA [1]

First Structural Characterization

The first structure of CBM41 was revealed in the x-ray crystal structure of full-length pullulanase from Klebsiella pneumoniae [4].

References

1. Lammerts van Bueren A, Finn R, Ausió J, and Boraston AB. (2004). Alpha-glucan recognition by a new family of carbohydrate-binding modules found primarily in bacterial pathogens. Biochemistry. 2004;43(49):15633-42. DOI:10.1021/bi048215z |
2. van Bueren AL and Boraston AB. (2007). The structural basis of alpha-glucan recognition by a family 41 carbohydrate-binding module from Thermotoga maritima. J Mol Biol. 2007;365(3):555-60. DOI:10.1016/j.jmb.2006.10.018 |
3. Lammerts van Bueren A, Ficko-Blean E, Pluvinage B, Hehemann JH, Higgins MA, Deng L, Ogunniyi AD, Stroeher UH, El Warry N, Burke RD, Czjzek M, Paton JC, Vocadlo DJ, and Boraston AB. (2011). The conformation and function of a multimodular glycogen-degrading pneumococcal virulence factor. Structure. 2011;19(5):640-51. DOI:10.1016/j.str.2011.03.001 |
4. Mikami B, Iwamoto H, Malle D, Yoon HJ, Demirkan-Sarikaya E, Mezaki Y, and Katsuya Y. (2006). Crystal structure of pullulanase: evidence for parallel binding of oligosaccharides in the active site. J Mol Biol. 2006;359(3):690-707. DOI:10.1016/j.jmb.2006.03.058 |
5. Gilbert HJ, Knox JP, and Boraston AB. (2013). Advances in understanding the molecular basis of plant cell wall polysaccharide recognition by carbohydrate-binding modules. Curr Opin Struct Biol. 2013;23(5):669-77. DOI:10.1016/j.sbi.2013.05.005 |

6. Davies, G.J. and Sinnott, M.L. (2008) Sorting the diverse: the sequence-based classifications of carbohydrate-active enzymes. Biochem. J. (BJ Classic Paper, online only). DOI: 10.1042/BJ20080382

7. Boraston AB, Bolam DN, Gilbert HJ, and Davies GJ. (2004). Carbohydrate-binding modules: fine-tuning polysaccharide recognition. Biochem J. 2004;382(Pt 3):769-81. DOI:10.1042/BJ20040892 |
8. Hashimoto H (2006). Recent structural studies of carbohydrate-binding modules. Cell Mol Life Sci. 2006;63(24):2954-67. DOI:10.1007/s00018-006-6195-3 |
9. Shoseyov O, Shani Z, and Levy I. (2006). Carbohydrate binding modules: biochemical properties and novel applications. Microbiol Mol Biol Rev. 2006;70(2):283-95. DOI:10.1128/MMBR.00028-05 |
10. Guillén D, Sánchez S, and Rodríguez-Sanoja R. (2010). Carbohydrate-binding domains: multiplicity of biological roles. Appl Microbiol Biotechnol. 2010;85(5):1241-9. DOI:10.1007/s00253-009-2331-y |