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The amylase inhibitor montbretin A reveals a new glycosidase inhibition motif

Nature Chemical Biology volume 11, pages 691–696 (2015)Cite this article

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Abstract

The complex plant flavonol glycoside montbretin A is a potent (Ki = 8 nM) and specific inhibitor of human pancreatic α-amylase with potential as a therapeutic for diabetes and obesity. Controlled degradation studies on montbretin A, coupled with inhibition analyses, identified an essential high-affinity core structure comprising the myricetin and caffeic acid moieties linked via a disaccharide. X-ray structural analyses of the montbretin A–human α-amylase complex confirmed the importance of this core structure and revealed a novel mode of glycosidase inhibition wherein internal π-stacking interactions between the myricetin and caffeic acid organize their ring hydroxyls for optimal hydrogen bonding to the α-amylase catalytic residues D197 and E233. This novel inhibitory motif can be reproduced in a greatly simplified analog, offering potential for new strategies for glycosidase inhibition and therapeutic development.

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Figure 1: Characterization of MbA functionalities.
Figure 2: The binding mode of MbA.
Figure 3: Pre-formed characteristics of MbA structure in solution.
Figure 4: Chemical structure of mini-MbA analog inhibitor 10.

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Acknowledgements

We thank R. Maurus for helpful discussions regarding structural analyses and E. Goddard-Borger for advice on synthesis. We thank E. Kwan for preparation of HPA, T. Hill for isolation of human gut bacterial amylases, C. Hii, P. Lincez and K. Liu for additional technical assistance, H. Flint for providing clones of the human gut bacterial amylases and D.R. Rose (University of Waterloo) for supplies of ntMGAM, ctMGAM and ctSI. This work was supported by Operating and Proof of Principle grants from the Canadian Institutes of Health Research (CIHR; ref. nos. (FRN) 111082, 200704PPP (to G.D.B. and S.G.W.)), by an operating grant from the Natural Sciences and Engineering Research Council (NSERC) of Canada to R.J.A. and through the CDRD-Pfizer Innovation Fund. L.K.W. was supported by an Alexander Graham Bell Canada Graduate Scholarship (NSERC). S.G.W. is supported by a Tier 1 Canada Research Chair. Portions of this research were carried out at the Stanford Synchrotron Radiation Lightsource (SSRL), SLAC National Accelerator Laboratory, which is supported by the US Department of Energy (DOE), Office of Science, Office of Basic Energy Sciences under contract no. DE-AC02-76SF00515. The SSRL Structural Molecular Biology Program is supported by the DOE Office of Biological and Environmental Research and by the National Institutes of Health (NIH), National Institute of General Medical Sciences (NIGMS) (including P41GM103393). The contents of this publication are solely the responsibility of the authors and do not necessarily represent the official views of NIGMS or NIH. The graphics program PyMOL (ref. 21) was used in part to generate figures.

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Author notes

  1. Leslie K Williams and Xiaohua Zhang: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of British Columbia, Vancouver, British Columbia, Canada

    Leslie K Williams, Sami Caner, Nham T Nguyen, Stephen G Withers & Gary D Brayer

  2. Department of Chemistry, Faculty of Science, University of British Columbia, Vancouver, British Columbia, Canada

    Xiaohua Zhang, Jacqueline Wicki, David E Williams, Raymond J Andersen & Stephen G Withers

  3. Genome Sciences and Technology Graduate Programme, University of British Columbia, Vancouver, British Columbia, Canada

    Christina Tysoe

  4. Department of Earth, Ocean and Atmospheric Sciences, University of British Columbia, Vancouver, British Columbia, Canada

    David E Williams & Raymond J Andersen

  5. Centre for Drug Research and Development, University of British Columbia, Vancouver, British Columbia, Canada

    John Coleman

  6. Faculty of Pharmaceutical Sciences, University of British Columbia, Vancouver, British Columbia, Canada

    John H McNeill & Violet Yuen

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Contributions

N.T.N. crystallized the MbA–HPA complex, and L.K.W. and S.C. solved and analyzed its structure; X.Z. performed the degradation studies of MbA and kinetic studies thereon; C.T. synthesized inhibitor 10 and performed kinetic studies thereon; J.W. performed kinetic studies on MbA variants; D.E.W. performed the ROESY NMR studies on MbA; J.C., V.Y., J.M., R.J.A., S.G.W. and G.D.B. designed and supervised the project. Writing was primarily done by V.Y., G.D.B., L.K.W. and S.G.W. with contributions from all.

Corresponding author

Correspondence to Stephen G Withers.

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Competing interests

The University of British Columbia holds a patent on the use of MbA as a blood glucose controlling agent with R.J.A., G.D.B. and S.G.W. as inventors.

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Supplementary Results, Supplementary Tables 1–3, Supplementary Figures 1–5 and Supplementary Notes 1 and 2. (PDF 1922 kb)

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Williams, L., Zhang, X., Caner, S. et al. The amylase inhibitor montbretin A reveals a new glycosidase inhibition motif. Nat Chem Biol 11, 691–696 (2015). https://doi.org/10.1038/nchembio.1865

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