title: Structural and mechanistic analysis of two prolyl endopeptidases: role of interdomain dynamics in catalysis and specificity.
abstract: Prolyl endopeptidases (PEPs) are a unique class of serine proteases with considerable therapeutic potential for the treatment of celiac sprue. The crystal structures of two didomain PEPs have been solved in alternative configurations, thereby providing insights into the mode of action of these enzymes. The structure of the Sphingomonas capsulata PEP, solved and refined to 1.8-A resolution, revealed an open configuration of the active site. In contrast, the inhibitor-bound PEP from Myxococcus xanthus was crystallized (1.5-A resolution) in a closed form. Comparative analysis of the two structures highlights a critical role for the domain interface in regulating interdomain dynamics and substrate specificity. Structure-based mutagenesis of the M. xanthus PEP confirms an important role for several interfacial residues. A salt bridge between Arg-572 and Asp-196/Glu-197 appears to act as a latch for opening or closing the didomain enzyme, and Arg-572 and Ile-575 may also help secure the incoming peptide substrate to the open form of the enzyme. Arg-618 and Asp-145 are responsible for anchoring the invariant proline residue in the active site of this postproline-cleaving enzyme. A model is proposed for the docking of a representative substrate PQPQLPYPQPQLP in the active site, where the N-terminal substrate residues interact extensively with the catalytic domain, and the C-terminal residues stretch into the propeller domain. Given the promise of the M. xanthus PEP as an oral therapeutic enzyme for treating celiac sprue, our results provide a strong foundation for further optimization of the PEP's clinically useful features.
authors.affiliation: Department of Chemical Engineering, Stanford University, Stanford, CA 94305, USA.
authors.forename: Lu
authors.initials: L
authors.lastname: Shan
authors.forename: Irimpan I
authors.initials: II
authors.lastname: Mathews
authors.forename: Chaitan
authors.initials: C
authors.lastname: Khosla
mesh_terms: D001665:Binding Sites; D020134:Catalytic Domain; D016941:Myxococcus xanthus / Q000201:enzymology*; D000085822:Prolyl Oligopeptidases; D017433:Protein Structure, Secondary; D012697:Serine Endopeptidases / Q000737:chemistry*; D020578:Sphingomonas / Q000201:enzymology*; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: D012697:Serine Endopeptidases; D000085822:Prolyl Oligopeptidases
references.citation: Protein Sci. 2004 Feb;13(2):412-21
references.pmid: 14718659
references.citation: Cell Mol Life Sci. 2002 Feb;59(2):349-62
references.pmid: 11915948
references.citation: Acta Crystallogr D Biol Crystallogr. 1998 Sep 1;54(Pt 5):905-21
references.pmid: 9757107
references.citation: Eur J Pharmacol. 1991 Nov 26;205(2):157-63
references.pmid: 1812006
references.citation: Acta Crystallogr A. 1991 Mar 1;47 ( Pt 2):110-9
references.pmid: 2025413
references.citation: Biochem J. 2004 Oct 15;383(Pt 2):311-8
references.pmid: 15245330
references.citation: J Biol Chem. 2001 Jan 12;276(2):1262-6
references.pmid: 11031266
references.citation: EMBO Rep. 2000 Sep;1(3):277-81
references.pmid: 11256612
references.citation: J Mol Biol. 2004 Jul 9;340(3):627-37
references.pmid: 15210359
references.citation: Biochemistry. 1991 Feb 26;30(8):2195-203
references.pmid: 1900195
references.citation: Structure. 2003 Aug;11(8):947-59
references.pmid: 12906826
references.citation: Arch Biochem Biophys. 1998 Oct 1;358(1):141-8
references.pmid: 9750174
references.citation: Biochem J. 1992 May 1;283 ( Pt 3):647-8
references.pmid: 1590752
references.citation: Am J Physiol Gastrointest Liver Physiol. 2002 Oct;283(4):G996-G1003
references.pmid: 12223360
references.citation: Curr Opin Struct Biol. 1998 Oct;8(5):606-11
references.pmid: 9818265
references.citation: J Biochem. 1991 Dec;110(6):873-8
references.pmid: 1840588
references.citation: Structure. 2000 May 15;8(5):R105-10
references.pmid: 10801499
references.citation: Adv Protein Chem. 1968;23:283-438
references.pmid: 4882249
references.citation: Biol Pharm Bull. 1997 Oct;20(10):1047-50
references.pmid: 9353562
references.citation: J Biol Chem. 1980 May 25;255(10):4786-92
references.pmid: 6768725
references.citation: J Biol Chem. 2001 Jun 1;276(22):19310-7
references.pmid: 11278687
references.citation: Acta Crystallogr A. 2003 May;59(Pt 3):228-34
references.pmid: 12714773
references.citation: J Pharmacol Exp Ther. 2005 Jan;312(1):19-26
references.pmid: 15358813
references.citation: Biochemistry. 1992 Nov 10;31(44):10769-73
references.pmid: 1420194
references.citation: J Mol Biol. 1993 Jun 20;231(4):1049-67
references.pmid: 8515464
references.citation: Cell. 1998 Jul 24;94(2):161-70
references.pmid: 9695945
references.citation: Acta Crystallogr D Biol Crystallogr. 1999 Apr;55(Pt 4):849-61
references.pmid: 10089316
references.citation: FEBS Lett. 1988 Jul 18;234(2):251-6
references.pmid: 3292288
references.citation: Science. 2002 Sep 27;297(5590):2275-9
references.pmid: 12351792