Carbohydrate Binding Module Family 14

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• Author: ^^^Eva Madland^^^
• Responsible Curator: ^^^Elizabeth Ficko-Blean^^^

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CAZy DB link

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

Ligand specificities

Family 14 CBMs are modules composed of approximately 70 residues. These modules have been reported to be associated with chitinases [1] and as chitin-binding lectins e.g. an effector protein from the tomato pathogen Pseudoercospora fuligena or Cladosporium fulvum [2, 3], as an antimicrobial lectin-like protein (tachycitin) from horseshoe crab haemocytes [4] and in peritrophic matrix proteins from Malaria vector Anopheles gambia [5]. Members of CBM14 have been shown to bind chitin [5, 6, 7] and chitooligomers [3, 8]. Binding to 50 % acetylated hyaluronan has also been demonstrated [8].

The ligand binding affinities have been quantified for two CBM14 members. The interaction between a human chitotriosidase-1 (CHIT1, characterized as a glycoside hydrolase family 18 (GH18)) CBM14 and (GlcNAc)₃ have been investigated with NMR titration experiments and isothermal titration calorimetry (ITC) displaying a relatively weak interaction of K_d 9.9 ± 0.8 mM [8] and K_d 3.1 ± 0.2 mM [7], respectively.

Interaction studies have also been performed for CBM14 in the fungal tomato pathogen C. fulvum and (GlcNAc)₆. Here, the binding properties were measured using ITC yielding a K_d of 6.7 ± 1.5 µM [3].

Structural Features

The structures of CBM14 members have a hevein-like fold commonly made up by a central β-sheet (three anti-parallel β-strands) linked to a small β-sheet (two anti-parallel β-strands) by two aromatic residues. The CBM14 members from tomato pathogens also have an N-terminal α-helix and an extended loop connecting -strands 2 and 3, as well as a C-terminal helical turn [2, 3]. The latter is also present in tachycitin [9]. In addtion, the CBM14 structures have been reported to contain 3-4 disulfide bridges which also serve as a stabilizing effect on this unique fold [1, 3, 9].

With this hevein-like fold and lectin properties, CBM14 is characterized as a Type C CBM [10]. This is in line with the ligand-binding feature seen for the CBM14 associated with CHIT1 (CBM14_CHIT1). The binding site is composed of a tryptophan and an asparagine (Trp465 and Asn466) at the C-terminus of this module [8]. These residues create a platform-like binding surface commonly seen in Type A CBMs and could be the reason for why CBM14_CHIT1 also is able to bind crystalline chitin. In addition to Trp465 and Asn466, a leucine (Leu454) have been reported [7] to be indirectly involved in binding as it contributes to maintaining a correct orientation for Trp465. Binding between CBM14_CHIT1 and chitotriose is likely to occur through CH-π stacking between the side-chain of Trp465 and the middle pyranose ring and hydrogen bonds between the side-chain of Asn466 and the hydrophilic non-reducing end [7].

CBM14 from the tomato pathogen C. fulvum (CfCBM14) was the first of these modules to be co-crystallized with (GlcNAc)₆ [3]. Although the fold of CfCBM14 is the same as CBM14_CHIT1 and other structurally characterized CBM14 members, the residues responsible for binding differ. Binding of (GlcNAc)₆ is mediated by Lys49, Cys50, Met51, Pro53, Lys99, Trp100, Cys101, Asp102 and Tyr103 positioned along a shallow trench in the longitudinal axis of CfCBM14. Mutational studies showed that binding to (GlcNAc)₆ was abolished by mutation of Trp100 and Asp102. The main interaction is reported to be a CH-π stacking between Trp100 and GlcNAc-5 aided by Met51 and Pro53 which are both in van der Waal distances to GlcNAc-1 and GlcNAc-3, respectively [3]. Although equivalents to Trp100, Asp102 and Tyr103 have been reported to be important for chitin-binding in CBM14 from tomato pathogen P. fuligena (PfCBM14) (equivalent residues: Trp94, Asp96 and Tyr97) [2], the extensive hydrogen bond network could point to a different binding mechanism for CfCBM14 that is more reminiscent of a Type B CBM.

Functionalities

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

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First Structural Characterization

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References

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10. 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 |