Abstract
Solution-state nuclear magnetic resonance (NMR) spectroscopy can be used to monitor protein-carbohydrate interactions. Two-dimensional 1H-15N heteronuclear single quantum coherence (HSQC)-based techniques described in this chapter can be used quickly and effectively to screen a set of possible carbohydrate binding partners, to quantify the dissociation constant (K d) of any identified interactions, and to map the carbohydrate binding site on the structure of the protein. Here, we describe the titration of a family 32 carbohydrate binding module from Clostridium perfringens (CpCBM32) with the monosaccharide N-acetylgalactosamine (GalNAc), in which we calculate the apparent dissociation of the interaction, and map the GalNAc binding site onto the structure of CpCBM32.
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References
Zuiderweg ER (2002) Mapping protein-protein interactions in solution by NMR spectroscopy. Biochemistry 41(1):1–7
Haselhorst T, Lamerz AC, Itzstein M (2009) Saturation transfer difference NMR spectroscopy as a technique to investigate protein-carbohydrate interactions in solution. Methods Mol Biol 534:375–386
Johnson MA, Pinto BM (2004) NMR spectroscopic and molecular modeling studies of protein-carbohydrate and protein-peptide interactions. Carbohydr Res 339(5):907–928
Kay LE, Keifer P, Saarinen T (1992) Pure absorption gradient enhanced heteronuclear single quantum correlation spectroscopy with improved sensitivity. J Am Chem Soc 114:10663–10665
Lian LY et al (1994) Protein-ligand interactions: exchange processes and determination of ligand conformation and protein-ligand contacts. Methods Enzymol 239:657–700
Johnson BA (2004) Using NMRView to visualize and analyze the NMR spectra of macromolecules. Methods Mol Biol 278:313–352
Vranken WF et al (2005) The CCPN data model for NMR spectroscopy: development of a software pipeline. Proteins 59(4):687–696
Grondin JM et al (2014) An unusual mode of galactose recognition by a family 32 carbohydrate-binding module. J Mol Biol 426(4):869–880
Viegas A et al (2013) Solution structure, dynamics and binding studies of a family 11 carbohydrate-binding module from Clostridium thermocellum (CtCBM11). Biochem J 451(2):289–300
Koay A et al (2007) Oligosaccharide recognition and binding to the carbohydrate binding module of AMP-activated protein kinase. FEBS Lett 581(26):5055–5059
Ohnuma T et al (2008) LysM domains from Pteris ryukyuensis chitinase-A: a stability study and characterization of the chitin-binding site. J Biol Chem 283(8):5178–5187
Boraston AB et al (2004) Carbohydrate-binding modules: fine-tuning polysaccharide recognition. Biochem J 382(Pt 3):769–781
Boraston AB, Ficko-Blean E, Healey M (2007) Carbohydrate recognition by a large sialidase toxin from Clostridium perfringens. Biochemistry 46(40):11352–11360
Ficko-Blean E, Boraston AB (2009) N-acetylglucosamine recognition by a family 32 carbohydrate-binding module from Clostridium perfringens NagH. J Mol Biol 390(2):208–220
Ficko-Blean E et al (2012) Carbohydrate recognition by an architecturally complex alpha-N-acetylglucosaminidase from Clostridium perfringens. PLoS One 7(3):e33524
Ficko-Blean E, Boraston AB (2006) The interaction of a carbohydrate-binding module from a Clostridium perfringens N-acetyl-beta-hexosaminidase with its carbohydrate receptor. J Biol Chem 281(49):37748–37757
Wilkins MR et al (1999) Protein identification and analysis tools in the ExPASy server. Methods Mol Biol 112:531–552
Shekels LL et al (1998) Cloning and characterization of mouse intestinal MUC3 mucin: 3' sequence contains epidermal-growth-factor-like domains. Biochem J 330(Pt 3):1301–1308
Delaglio F et al (1995) NMRPipe: a multidimensional spectral processing system based on UNIX pipes. J Biomol NMR 6(3):277–293
DeLano WL (2002) The PyMOL molecular graphics system, Version 1.8 Schrödinger, LLC
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Grondin, J.M., Langelaan, D.N., Smith, S.P. (2017). Characterization of Protein-Carbohydrate Interactions by NMR Spectroscopy. In: Abbott, D., Lammerts van Bueren, A. (eds) Protein-Carbohydrate Interactions. Methods in Molecular Biology, vol 1588. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6899-2_11
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DOI: https://doi.org/10.1007/978-1-4939-6899-2_11
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