Conformational response to ligand binding in phosphomannomutase2: insights into inborn glycosylation disorder

The most common glycosylation disorder is caused by mutations in the gene encoding phosphomannomutase2, producing a disease still without a cure. Phosphomannomutase2, a homodimer in which each chain is composed of two domains, requires a bisphosphate sugar (either mannose or glucose) as activator, o...

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Bibliographic Details
Published in:	The Journal of biological chemistry 2014-12, Vol.289 (50), p.34900-34910
Main Authors:	Andreotti, Giuseppina, Cabeza de Vaca, Israel, Poziello, Angelita, Monti, Maria Chiara, Guallar, Victor, Cubellis, Maria Vittoria
Format:	Article
Language:	eng
Subjects:	Amino Acid Sequence Animals Glucose-6-Phosphate - analogs & derivatives Glucose-6-Phosphate - metabolism Glycosylation Humans Ligands Metabolism, Inborn Errors - enzymology Models, Molecular Molecular Biophysics Molecular Sequence Data Mutation Peptide Hydrolases - metabolism Phosphotransferases (Phosphomutases) - chemistry Phosphotransferases (Phosphomutases) - genetics Phosphotransferases (Phosphomutases) - metabolism Protein Binding Protein Conformation Protein Unfolding Proteolysis Temperature
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Summary:	The most common glycosylation disorder is caused by mutations in the gene encoding phosphomannomutase2, producing a disease still without a cure. Phosphomannomutase2, a homodimer in which each chain is composed of two domains, requires a bisphosphate sugar (either mannose or glucose) as activator, opening a possible drug design path for therapeutic purposes. The crystal structure of human phosphomannomutase2, however, lacks bound substrate and a key active site loop. To speed up drug discovery, we present here the first structural model of a bisphosphate substrate bound to human phosphomannomutase2. Taking advantage of recent developments in all-atom simulation techniques in combination with limited and site-directed proteolysis, we demonstrated that α-glucose 1,6-bisphosphate can adopt two low energy orientations as required for catalysis. Upon ligand binding, the two domains come close, making the protein more compact, in analogy to the enzyme in the crystals from Leishmania mexicana. Moreover, proteolysis was also carried out on two common mutants, R141H and F119L. It was an unexpected finding that the mutant most frequently found in patients, R141H, although inactive, does bind α-glucose 1,6-bisphosphate and changes conformation.
ISSN:	0021-9258 1083-351X