Laforin is a human dual-specificity phosphatase (DSP) involved in glycogen metabolism regulation containing a carbohydrate-binding module (CBM). Mutations in the gene coding for laforin are responsible for the development of Lafora disease, a progressive fatal myoclonus epilepsy with early onset, characterized by the intracellular deposition of abnormally branched, hyperphosphorylated insoluble glycogen-like polymers, called Lafora bodies. Despite the known importance of the CBM domain of laforin in the regulation of glycogen metabolism, the molecular mechanism of laforin–glycogen interaction is still poorly understood. Recently, the structure of laforin with bound maltohexaose was determined and despite the importance of such breakthrough, some molecular interaction details remained missing. We herein report a thorough biophysical characterization of laforin–carbohydrate interaction using soluble glycans. We demonstrated an increased preference of laforin for the interaction with glycans with higher order of polymerization and confirmed the importance of tryptophan residues for glycan interaction. Moreover, and in line with what has been described for other CBMs and lectins, our results confirmed that laforin–glycan interactions occur with a favourable enthalpic contribution counter-balanced by an unfavourable entropic contribution. The analysis of laforin–glycan interaction through the glycan side by saturation transfer difference (STD)–NMR has shown that the CBM-binding site can accommodate between 5 and 6 sugar units, which is in line with the recently obtained crystal structure of laforin. Overall, the work in the present study complements the structural characterization of laforin and sheds light on the molecular mechanism of laforin–glycan interaction, which is a pivotal requisite to understand the physiological and pathological roles of laforin.
Biophysical characterization of laforin–carbohydrate interaction
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David M. Dias, Joana Furtado, Emeric Wasielewski, Rui Cruz, Bernard Costello, Lindsay Cole, Tiago Q. Faria, Philipp Baaske, Rui M. M. Brito, Alessio Ciulli, Isaura Simões, Sandra Macedo-Ribeiro, Carlos Faro, Carlos F. G. C. Geraldes, Pedro Castanheira; Biophysical characterization of laforin–carbohydrate interaction. Biochem J 1 February 2016; 473 (3): 335–345. doi: https://doi.org/10.1042/BJ20141555
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