Abstract
The three-dimensional structures of pepsin inhibitor-3 (PI-3) from Ascaris suum and of the complex between PI-3 and porcine pepsin at 1.75 Å and 2.45 Å resolution, respectively, have revealed the mechanism of aspartic protease inhibition by this unique inhibitor. PI-3 has a new fold consisting of two domains, each comprising an antiparallel β-sheet flanked by an α-helix. In the enzyme–inhibitor complex, the N-terminal β-strand of PI-3 pairs with one strand of the ‘active site flap’ (residues 70–82) of pepsin, thus forming an eight-stranded β-sheet that spans the two proteins. PI-3 has a novel mode of inhibition, using its N-terminal residues to occupy and therefore block the first three binding pockets in pepsin for substrate residues C-terminal to the scissile bond (S1′–S3′). The molecular structure of the pepsin–PI-3 complex suggests new avenues for the rational design of proteinaceous aspartic proteinase inhibitors.
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Acknowledgements
We thank L. Swenson for assisting with the purification of porcine pepsin; M. Carpenter for performing N-terminal sequencing and amino acid composition determinations; L. Burke for performing mass spectrometry analysis; M. Fraser, B. Mark, W. Wolodko and the staff of BioCARS for assistance with the MAD X-ray data collection; P. Roversi, E. Blanc and G. Bricogne for improving electron density maps of the pepsin–PI-3 complex with a developmental version of BUSTER-TNT; and N. Khazanovich Bernstein and M. Fujinaga for helpful discussions. Use of the Advanced Photon Source was supported by the US Department of Energy, Basic Energy Sciences, Office of Science. Use of BioCARS Sector 14 was supported by the National Institutes of Health, National Center for Research Resources. This work was supported by grants from the Medical Research Council of Canada (MRC) to M.N.G.J. and from the National Institutes of Health to B.M.D. K.K.S.N. was supported by a post-doctoral fellowship from the MRC and a research allowance supplement from the Alberta Heritage Fund for Medical Research. J.F.W.P. was supported by a Killam post-doctoral fellowship at the University of Alberta.
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Ng, K., Petersen, J., Cherney, M. et al. Structural basis for the inhibition of porcine pepsin by Ascaris pepsin inhibitor-3 . Nat Struct Mol Biol 7, 653–657 (2000). https://doi.org/10.1038/77950
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DOI: https://doi.org/10.1038/77950
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