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Biological Chemistry

Editor-in-Chief: Brüne, Bernhard

Editorial Board Member: Buchner, Johannes / Ludwig, Stephan / Sies, Helmut / Turk, Boris / Wittinghofer, Alfred

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Trypsin inhibition by macrocyclic and open-chain variants of the squash inhibitor MCoTI-II

Olga Avrutina1 / Hans-Ulrich Schmoldt2 / Dusica Gabrijelcic-Geiger3 / Dung Le Nguyen4 / Christian P. Sommerhoff5 / Ulf Diederichsen6 / Harald Kolmar7

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Corresponding author

Citation Information: Biological Chemistry. Volume 386, Issue 12, Pages 1301–1306, ISSN (Online) 1437-4315, ISSN (Print) 1431-6730, DOI: 10.1515/BC.2005.148, December 2005

Publication History

Received:
June 13, 2005
Accepted:
September 14, 2005
Published Online:
2005-12-09

Abstract

MCoTI-I and MCoTI-II from the seeds of Momordica cochinchinensis are inhibitors of trypsin-like proteases and the only known members of the large family of squash inhibitors that are cyclic and contain an additional loop connecting the amino- and the carboxy-terminus. To investigate the contribution of macrocycle formation to biological activity, we synthesized a set of open-chain variants of MCoTI-II that lack the cyclization loop and contain various natural and non-natural amino acid substitutions in the reactive-site loop. Upon replacement of P1 lysine residue #10 within the open-chain variant of MCoTI-II by the non-natural isosteric nucleo amino acid AlaG [β-(guanin-9-yl)-L-alanine], a conformationally restricted arginine mimetic, residual inhibitory activity was detected, albeit reduced by four orders of magnitude. While the cyclic inhibitors MCoTI-I and MCoTI-II were found to be very potent trypsin inhibitors, with picomolar inhibition constants, the open-chain variants displayed an approximately 10-fold lower affinity. These data suggest that the formation of a circular backbone in the MCoTI squash inhibitors results in enhanced affinity and therefore is a determinant of biological activity.

Keywords: cyclic squash inhibitors; cystine-knot microproteins; nucleic amino acids; trypsin inhibition

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