Abstract
Gaucher disease results from mutations in the lysosomal enzyme acid β-glucosidase (GCase)1. Although enzyme replacement therapy has improved the health of some affected individuals, such as those with the prevalent N370S mutation, oral treatment with pharmacological chaperones may be therapeutic in a wider range of tissue compartments by restoring sufficient activity of endogenous mutant GCase2. Here we demonstrate that isofagomine (IFG, 1) binds to the GCase active site, and both increases GCase activity in cell lysates and restores lysosomal trafficking in cells containing N370S mutant GCase. We also compare the crystal structures of IFG-bound GCase at low pH with those of glycerol-bound GCase at low pH and apo-GCase at neutral pH. Our data indicate that IFG induces active GCase, which is secured by interactions with Asn370. The design of small molecules that stabilize substrate-bound conformations of mutant proteins may be a general therapeutic strategy for diseases caused by protein misfolding and mistrafficking.
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Acknowledgements
This work was supported in part by a US National Institutes of Health (NIH) National Research Service Award to R.L.L., and by a grant from the American Parkinson Disease Association (to G.A.P.). M.G.S. is supported by the US NIH National Institute of Neurologic Disorders and Stroke (Udall Center for Excellence in Parkinson Disease Research). We thank M. Wolfe for use of laboratory equipment, R. Sanishvili for assistance with data collection and processing, W. Novak for assistance with in silico screening, and G. Grabowski, S. Kornfeld, V. Cullen, H. Do, E. Benjamin, M. Toth, G. Lee and K. Valezano for helpful discussions. We are also thankful for the support and pioneering efforts of J. Fan and K. Chang in using pharmacological chaperones for the rescue of misfolded proteins. GM/CA-CAT is funded by the US National Cancer Institute and the US National Institute of General Medical Sciences. Use of the Advanced Photon Source was supported by the US Department of Energy. We thank J.-Q. Fan (Mount Sinai School of Medicine) for the rabbit polyclonal anti-GCase.
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R.L.L. conducted the crystallographic studies and prepared the manuscript. A.C.P., C.W.P. and R.K. developed protocols and generated data for GCase inhibition, enhancement and immunofluorescence studies. M.G.S. aided in the purchase of Cerezyme and the interpretation of results, and commented on the manuscript. B.A.W. and P.H. conceived of and directed the project. D.R. and G.A.P. provided overall guidance and direction in experimentation, interpretation and manuscript preparation.
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P.H., B.A.W., A.C.P., C.W.P. and R.K. are employees of Amicus Therapeutics, developing pharmacological chaperones for Gaucher and other diseases. M.G.S. and G.A.P. are advisors to Amicus Therapeutics. M.G.S. is the co-recipient of a translational research grant from Genzyme Corporation (not related to this work).
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Data collection and refinement statistics. (PDF 31 kb)
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Lieberman, R., Wustman, B., Huertas, P. et al. Structure of acid β-glucosidase with pharmacological chaperone provides insight into Gaucher disease. Nat Chem Biol 3, 101–107 (2007). https://doi.org/10.1038/nchembio850
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DOI: https://doi.org/10.1038/nchembio850
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