New to the CAZy classification? Read this first.
Want to learn more about CAZypedia? Read the CAZypedia 10th anniversary article in Glycobiology.
Carbohydrate Binding Module Family 14
This page is currently under construction. This means that the Responsible Curator has deemed that the page's content is not quite up to CAZypedia's standards for full public consumption. All information should be considered to be under revision and may be subject to major changes.
|CAZy DB link|
Family 14 CBMs are modules composed of approximately 70 residues. These modules have been reported to be associated with chitinases  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 hemocytes  and in peritrophic matrix proteins from Malaria vector Anopheles gambia . 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 .
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 Kd 9.9 ± 0.8 mM  and Kd 3.1 ± 0.2 mM , 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 Kd of 6.7 ± 1.5 µM .
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 . 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 . This is in line with the ligand-binding feature seen for the CBM14 associated with CHIT1 (CBM14CHIT1). The binding site is composed of a tryptophan and an asparagine (Trp465 and Asn466) at the C-terminus of this module  (Figure 1A). These residues create a platform-like binding surface (Figure 1B) commonly seen in Type A CBMs and could be the reason for why CBM14CHIT1 also is able to bind crystalline chitin. In addition to Trp465 and Asn466, a leucine (Leu454) have been reported  to be indirectly involved in binding as it contributes to maintaining a correct orientation for Trp465. Binding between CBM14CHIT1 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 .
CBM14 from the tomato pathogen C. fulvum (CfCBM14) was the first of these modules to be co-crystallized with (GlcNAc)6 (PDB ID: 6BN0). Although the fold of CfCBM14 is the same as CBM14CHIT1 (PDB ID: 5HBF and 6SO0) 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 (Figure 1C and D). 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 . 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) , the extensive hydrogen bond network could point to a different binding mechanism for CfCBM14 that is more reminiscent of a Type B CBM.
Several members of CBM14 are often encountered as chitin-binding lectins [2, 3, 4, 9, 11], while the members present in modular chitinases play a key role in targeting substrate. The most common associated modules are chitinases, e.g. GH18 from human chitotriosidase structurally determined together with CBM14CHIT1 (PDB ID: 5HBF).
Although some of the structural features described above differ between the lectin-CBM14 and CBM14CHIT1, the common denominator is the involvement of these modules in immune responses. CHIT1 is produced by macrophages and neutrophils [12, 13] and is utilized by innate immune response to combat chitin-containing pathogens . A similar response is found in the tomato pathogen C. fulvum. Here, the effector protein utilizes CBM14's ability to bind chitin in the fungal cell wall enabling protection from hydrolysis by plant-derived chitinases during infection [15, 16].
- First Identified
- CBM14 was first identified as an antimicrobial lectin-like protein with chitin binding ability in the hemocyte of horseshoe crab (Tachypleus tridentatus) and called tachycitin .
- First Structural Characterization
- The first three-dimensional structure was determined by NMR spectroscopy for the CBM14 module tachycitin (PDB ID: 1DQC) . The structure of the first carbohydrate-active enzyme associated CBM14 was determined by X-ray crystallography for the CBM14 of human chitotriosidase (PDB ID: 5HBF) .
- Fadel F, Zhao Y, Cousido-Siah A, Ruiz FX, Mitschler A, and Podjarny A. (2016). X-Ray Crystal Structure of the Full Length Human Chitotriosidase (CHIT1) Reveals Features of Its Chitin Binding Domain. PLoS One. 2016;11(4):e0154190. DOI:10.1371/journal.pone.0154190 |
- Kohler AC, Chen LH, Hurlburt N, Salvucci A, Schwessinger B, Fisher AJ, and Stergiopoulos I. (2016). Structural Analysis of an Avr4 Effector Ortholog Offers Insight into Chitin Binding and Recognition by the Cf-4 Receptor. Plant Cell. 2016;28(8):1945-65. DOI:10.1105/tpc.15.00893 |
- Hurlburt NK, Chen LH, Stergiopoulos I, and Fisher AJ. (2018). Structure of the Cladosporium fulvum Avr4 effector in complex with (GlcNAc)6 reveals the ligand-binding mechanism and uncouples its intrinsic function from recognition by the Cf-4 resistance protein. PLoS Pathog. 2018;14(8):e1007263. DOI:10.1371/journal.ppat.1007263 |
- Kawabata S, Nagayama R, Hirata M, Shigenaga T, Agarwala KL, Saito T, Cho J, Nakajima H, Takagi T, and Iwanaga S. (1996). Tachycitin, a small granular component in horseshoe crab hemocytes, is an antimicrobial protein with chitin-binding activity. J Biochem. 1996;120(6):1253-60. DOI:10.1093/oxfordjournals.jbchem.a021549 |
- Shen Z and Jacobs-Lorena M. (1998). A type I peritrophic matrix protein from the malaria vector Anopheles gambiae binds to chitin. Cloning, expression, and characterization. J Biol Chem. 1998;273(28):17665-70. DOI:10.1074/jbc.273.28.17665 |
- Vandevenne M, Campisi V, Freichels A, Gillard C, Gaspard G, Frère JM, Galleni M, and Filée P. (2011). Comparative functional analysis of the human macrophage chitotriosidase. Protein Sci. 2011;20(8):1451-63. DOI:10.1002/pro.676 |
- Madland E, Crasson O, Vandevenne M, Sørlie M, and Aachmann FL. (2019). NMR and Fluorescence Spectroscopies Reveal the Preorganized Binding Site in Family 14 Carbohydrate-Binding Module from Human Chitotriosidase. ACS Omega. 2019;4(26):21975-21984. DOI:10.1021/acsomega.9b03043 |
- Crasson O, Courtade G, Léonard RR, Aachmann FL, Legrand F, Parente R, Baurain D, Galleni M, Sørlie M, and Vandevenne M. (2017). Human Chitotriosidase: Catalytic Domain or Carbohydrate Binding Module, Who's Leading HCHT's Biological Function. Sci Rep. 2017;7(1):2768. DOI:10.1038/s41598-017-02382-z |
- Suetake T, Tsuda S, Kawabata S, Miura K, Iwanaga S, Hikichi K, Nitta K, and Kawano K. (2000). Chitin-binding proteins in invertebrates and plants comprise a common chitin-binding structural motif. J Biol Chem. 2000;275(24):17929-32. DOI:10.1074/jbc.C000184200 |
- 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 |
- van den Burg HA, Spronk CA, Boeren S, Kennedy MA, Vissers JP, Vuister GW, de Wit PJ, and Vervoort J. (2004). Binding of the AVR4 elicitor of Cladosporium fulvum to chitotriose units is facilitated by positive allosteric protein-protein interactions: the chitin-binding site of AVR4 represents a novel binding site on the folding scaffold shared between the invertebrate and the plant chitin-binding domain. J Biol Chem. 2004;279(16):16786-96. DOI:10.1074/jbc.M312594200 |
- Hollak CE, van Weely S, van Oers MH, and Aerts JM. (1994). Marked elevation of plasma chitotriosidase activity. A novel hallmark of Gaucher disease. J Clin Invest. 1994;93(3):1288-92. DOI:10.1172/JCI117084 |
- Kzhyshkowska J, Gratchev A, and Goerdt S. (2007). Human chitinases and chitinase-like proteins as indicators for inflammation and cancer. Biomark Insights. 2007;2:128-46. | Google Books | Open Library
- Gordon-Thomson C, Kumari A, Tomkins L, Holford P, Djordjevic JT, Wright LC, Sorrell TC, and Moore GP. (2009). Chitotriosidase and gene therapy for fungal infections. Cell Mol Life Sci. 2009;66(6):1116-25. DOI:10.1007/s00018-009-8765-7 |
- Joosten MH, Vogelsang R, Cozijnsen TJ, Verberne MC, and De Wit PJ. (1997). The biotrophic fungus Cladosporium fulvum circumvents Cf-4-mediated resistance by producing unstable AVR4 elicitors. Plant Cell. 1997;9(3):367-79. DOI:10.1105/tpc.9.3.367 |
- van den Burg HA, Harrison SJ, Joosten MH, Vervoort J, and de Wit PJ. (2006). Cladosporium fulvum Avr4 protects fungal cell walls against hydrolysis by plant chitinases accumulating during infection. Mol Plant Microbe Interact. 2006;19(12):1420-30. DOI:10.1094/MPMI-19-1420 |