title: Trypsin inhibition by macrocyclic and open-chain variants of the squash inhibitor MCoTI-II.
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 [beta-(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.
authors.affiliation: Institute for Organic and Biomolecular Chemistry, Georg-August-University, D-37077 Göttingen, Germany.
authors.forename: Olga
authors.initials: O
authors.lastname: Avrutina
authors.forename: Hans-Ulrich
authors.initials: HU
authors.lastname: Schmoldt
authors.forename: Dusica
authors.initials: D
authors.lastname: Gabrijelcic-Geiger
authors.forename: Dung
authors.initials: D
authors.lastname: Le Nguyen
authors.forename: Christian P
authors.initials: CP
authors.lastname: Sommerhoff
authors.forename: Ulf
authors.initials: U
authors.lastname: Diederichsen
authors.forename: Harald
authors.initials: H
authors.lastname: Kolmar
mesh_terms: D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D001120:Arginine / Q000737:chemistry; D002864:Chromogranins / Q000737:chemistry; D019662:Cucurbitaceae / Q000201:enzymology*; D003500:Cyclization; D047169:Cyclotides / Q000138:chemical synthesis* / Q000494:pharmacology; D008239:Lysine / Q000737:chemistry; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D011487:Protein Conformation; D014357:Trypsin / Q000378:metabolism*
chemical_list: D002864:Chromogranins; D047169:Cyclotides; C046560:beta-granins; C409923:trypsin inhibitor MCoTI-II; D001120:Arginine; D014357:Trypsin; D008239:Lysine; D000409:Alanine