ABSTRACT
With the aim of developing polyproline type II helix (PPII) secondary-structure mimetics for the modulation of prolin-rich-mediated protein-protein interactions, the novel diproline mimetic ProM-2 was designed by bridging the two pyrrolidine rings of a diproline (Pro-Pro) unit through a Z-vinylidene moiety. This scaffold, which closely resembles a section of a PPII helix, was then stereoselectively synthesized by exploiting a ruthenium-catalyzed ring-closing metathesis (RCM) as a late key step. The required vinylproline building blocks, that is, (R)-N-Boc-2-vinylproline (Boc=tert-butyloxycarbonyl) and (S,S)-5-vinylproline-tert-butyl ester, were prepared on a gram scale as pure stereoisomers. The difficult peptide coupling of the sterically demanding building blocks was achieved in good yield and without epimerization by using 2-(1H-7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HATU)/N,N-diisopropylethylamine (DIPEA). The RCM proceeded smoothly in the presence of the Grubbs II catalyst. Stereostructural assignments for several intermediates were secured by X-ray crystallography. As a proof of concept, it was shown that certain peptides containing ProM-2 exhibited improved (canonical) binding towards the Ena/VASP homology 1 (EVH1) domain as a relevant protein interaction target.
Subject(s)
Peptides/chemistry , Proteins/chemistry , Dipeptides/chemistry , Peptidomimetics , Protein Conformation , StereoisomerismABSTRACT
Small-molecule competitors of protein-protein interactions are urgently needed for functional analysis of large-scale genomics and proteomics data. Particularly abundant, yet so far undruggable, targets include domains specialized in recognizing proline-rich segments, including Src-homology 3 (SH3), WW, GYF, and Drosophila enabled (Ena)/vasodilator-stimulated phosphoprotein (VASP) homology 1 (EVH1) domains. Here, we present a modular strategy to obtain an extendable toolkit of chemical fragments (ProMs) designed to replace pairs of conserved prolines in recognition motifs. As proof-of-principle, we developed a small, selective, peptidomimetic inhibitor of Ena/VASP EVH1 domain interactions. Highly invasive MDA MB 231 breast-cancer cells treated with this ligand showed displacement of VASP from focal adhesions, as well as from the front of lamellipodia, and strongly reduced cell invasion. General applicability of our strategy is illustrated by the design of an ErbB4-derived ligand containing two ProM-1 fragments, targeting the yes-associated protein 1 (YAP1)-WW domain with a fivefold higher affinity.
Subject(s)
Proline-Rich Protein Domains , Protein Interaction Mapping , Animals , Cell Adhesion Molecules/chemistry , Cell Line, Tumor , Cell Membrane Permeability , Crystallography, X-Ray , Drosophila melanogaster/metabolism , Esterification , Fluorescent Antibody Technique , Humans , Kinetics , Ligands , Microfilament Proteins/chemistry , Models, Molecular , Molecular Weight , Peptides/chemistry , Phosphoproteins/chemistry , Protein Binding , Protein Structure, Tertiary , Pseudopodia , Stress Fibers/metabolism , Zyxin/chemistryABSTRACT
A practical and scalable synthesis of a Fmoc-protected tricyclic dipeptide mimetic (6), that is, a 1,4-diaza-tricyclo-[8.3.0(3,7)]-tridec-8-ene derivative resembling a rigidified di-L-proline in a polyproline type II (PPII) helix conformation, was developed. The strategy is based on a Ru-catalyzed ring-closing metathesis of a dipeptide (4) prepared by PyBOP coupling of cis-5-vinylproline tert-butylester (2) and trans-N-Boc-3-vinylproline (rac-3) followed by chromatographic diastereomer separation. Building block 2 was prepared from L-proline in six steps via electrochemical C5-methoxylation, cyanation and conversion of the nitrile into a vinyl substituent. Building block rac-3 was prepared in five steps exploiting a Cu-catalyzed 1,4-addition of vinyl-MgBr to a 2,3-dehydroproline derivative in the key step. In the course of the investigation subtle dependencies of protecting groups on the reactivity of the 2,3- and 2,5-disubstituted pyrrolidine derivatives were observed. The configuration and conformational preference of several intermediates were determined by X-ray crystallography. The developed synthesis allows the preparation of substantial amounts of 6, which will be used in the search for new small molecules for the modulation of protein-protein interactions involving proline-rich motifs (PRDs).
Subject(s)
Dipeptides/chemistry , Peptides/chemistry , Catalysis , Copper/chemistry , Crystallography, X-Ray , Dipeptides/chemical synthesis , Models, Molecular , Protein Conformation , Protein Interaction Domains and Motifs , StereoisomerismSubject(s)
Dipeptides/chemistry , Peptides/chemistry , Proline-Rich Protein Domains , Amino Acid Sequence , Ligands , Magnetic Resonance Spectroscopy , Peptides/chemical synthesis , Protein Binding , Protein Interaction Domains and Motifs , Protein Structure, Secondary , Protein Structure, Tertiary , StereoisomerismABSTRACT
In the title compound, K(+)·C(6)H(8)NO(4) (-), the K(+) cations have a coordination number of seven and are surrounded by four bidentate azinate anions. The methyl-ene groups of the anions are always directed towards the coordinated potassium cations. The N-C-C-C torsion angle is 101.2â (2)°. The orthogonal non-conjugated nature of the salt confirms the supposed geometry and reactivity of this compound.
