A Template‐Assembled Synthetic Protein Surface Mimetic of the von Willebrand Factor A1 domain Inhibits Botrocetin‐Induced Platelet Aggregation

Hauert, J; Fernández‐Carneado, Jimena; Michielin, Olivier; Mathieu, Sandrine; Grell, Daniel; Schapira, Marc; Spertini, Olivier; Mutter, Manfred; Tuchscherer, Gabriele; Kovacsovics, Tibor

doi:10.1002/cbic.200300826

Cited by 16 publications

(12 citation statements)

References 32 publications

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“…With improved understanding of how the different forces interact to influence the folded protein, it should be possible to direct the design of more complex caviteins with function. De novo proteins have been designed to mimic ion channels, [39] receptor binding proteins, [40] antiviral agents, [41] enzymes [42,43] and artificial redox systems. [43,44] Figure 11.…”

Section: Discussionmentioning

confidence: 99%

Optimal Attachment Position and Linker Length Promote Native‐like Character of Cavitand‐Based Template‐Assembled Synthetic Proteins (TASPs)

Seo¹,

Scott²,

Straus³

et al. 2007

Chemistry A European J

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We have designed, synthesised and characterised a series of template-assembled de novo four-helix bundles, each differing in the linker length between the template and the peptides. The helix is based on an earlier peptide sequence: EELLKKLEELLKKLG (first-generation sequence), which was designed to link the hydrophilic/hydrophobic interface of the helices. Increasing or decreasing the linker length by one glycine residue had a significant effect on the structure and properties of the template-assembled synthetic proteins (TASPs). Here, the effect of the linker length is further probed by linking the peptides closer to the hydrophobic face by using the second-generation sequence, AEELLKKLEELLKKG, in an effort to improve the packing between the helices and to better understand the helical bundles. The peptides were synthesised with 0-4 Gly linker residues and linked onto a cavitand template. The proteins were found to be alpha-helical, stable to guanidine hydrochloride (GuHCl) and to unfold cooperatively. However, their stabilities toward GuHCl, propensity to self-aggregate and structural specificity differed. The two-glycine variant of the second-generation series demonstrated the highest stability and most native-like character of all the mononeric TASPs in both the first- and second-generation series. The structural specificity of this two glycine variant is comparable to that of other known native-like de novo proteins. Molecular dynamics simulations showed that the two-glycine variant contains helices that are tilted with respect to the cavitand template and may account for its unique properties.

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Section: Discussionmentioning

confidence: 99%

Optimal Attachment Position and Linker Length Promote Native‐like Character of Cavitand‐Based Template‐Assembled Synthetic Proteins (TASPs)

Seo¹,

Scott²,

Straus³

et al. 2007

Chemistry A European J

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“…The use of RAFT molecules as scaffolds for the introduction of function by the grafting of surface residues present on native proteins is another area of major research interest in protein engineering and de novo design. Hauert et al designed a template‐assembled synthetic protein‐surface mimetic of the von Willebrand factor (WWF) A1 domain 119. The interaction of WWF with its platelet receptor GPIb‐IX complex mediates platelet adhesion.…”

Section: Applicationsmentioning

confidence: 99%

“…A chimeric 4‐helix bundle synthetic‐protein mimicking the surface of the A1 domain was prepared on a cyclic peptide scaffold by grafting 12 residues located on the α3 and α4 helices of the native protein. This synthetic construct was found to specifically inhibit botrocetin‐induced platelet aggregation and showed binding to both botrocetin and GPIbα 119…”

Section: Applicationsmentioning

confidence: 99%

Synthetic Peptide Templates for Molecular Recognition: Recent Advances and Applications

et al. 2006

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The creation of molecular systems that can mimic some of the properties of natural macromolecules is one of the major endeavors in contemporary protein chemistry. However, the construction of artificial proteins with predetermined structure and function is difficult on account of complex folding pathways. The use of topological peptide templates has been suggested to induce and stabilize defined secondary and tertiary structures. This is because the recent advances in the chemistry of coupling reagents, protecting groups, and solid-phase synthesis have made the chemical synthesis of peptides with conformationally controlled and complex structures feasible. Besides their use as structure-inducing devices, these peptide templates can also be utilized to construct novel structures with tailor-made functions. Herein, we present recent advances in the field of peptide-template-based approaches with particular emphasis on the demonstrated utility of this approach in molecular recognition, along with related applications.

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“…Various TASPs have been developed for the investigation of protein folding2 and peptide mimicry for drug design3 since Mutter and co‐workers pioneered the concept of TASP 1. In preceding works, a variety of templates were employed for TASPs, such as peptides,3a, d, e, 4, 5 saccharides,6 steroids,3c, 7 porphyrins,8 cavitands,2a, b, 9 and calixarenes 10. Although the structures of these TASPs have been analyzed with NMR, CD, and UV/Vis spectroscopy,2–10 these methodologies are difficult for the analysis of the stereostructure of TASPs at atomic resolution.…”

Section: Torsion Angles [°] In the Methylene Bridges Of The Calixarenmentioning

confidence: 99%

Calix[6]arene‐Based Template for X‐ray Crystallographic Analysis of Template‐Assembled Synthetic Proteins

Tsukamoto¹,

Ohishi²,

Maezaki³

et al. 2006

ChemBioChem

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Template-assembled synthetic proteins (TASPs) are artificial proteins in which peptides are covalently connected to a topological template in order to overcome potential misfolding by intermolecular folding.[1] Various TASPs have been developed for the investigation of protein folding [2] and peptide mimicry for drug design [3] since Mutter and co-workers pioneered the concept of TASP.[1] In preceding works, a variety of templates were employed for TASPs, such as peptides, [3a, d, e, 4, 5] saccharides, [6] steroids, [3c, 7] porphyrins, [8] cavitands, [2a, b, 9] and calixarenes.[10] Although the structures of these TASPs have been analyzed with NMR, CD, and UV/Vis spectroscopy, [2][3][4][5][6][7][8][9][10] these methodologies are difficult for the analysis of the stereostructure of TASPs at atomic resolution. X-ray crystallographic analysis is a useful method, [2a] but there are few reports on the X-ray analysis of TASPs.[11]Herein, we report the synthesis and X-ray analysis of p-tertbutylcalix[6]arene p-bromophenylalanine derivative 1. This novel topological template for TASPs facilitates phasing in Xray analysis through its bromine atoms.We employed the calixarene-type template 1, [12] which is readily synthesized from 1,3,5-trimethoxy p-tert-butylcalix[6]arene tricarboxylic acid 2, [13] for the following reasons: 1) calixarene derivatives show high crystallinity; 2) it has three tethers for assembly with peptides; 3) it has a flattened-cone conformation [14] which holds the peptide chains in the same direction; 4) it has the appropriate interval between carboxyl groups for the introduction of a-helices or b-sheets; 5) it facilitates phase calculation in X-ray crystallographic analysis since bromine atoms are introduced as a heavy-atom source. Conformational analyses of calix [4]-and -[6]arene amino acids or peptide derivatives have been reported. [15,16] The results indicated that intramolecular and/or intermolecular hydrogen bonds between amide functions affect their conformations and that the calixarene moieties have stably contained cone or flattenedcone conformations.To confirm its stereochemistry in the solid phase and validity as a template, X-ray crystallographic analysis of 1 was performed. A crystal of 1 was obtained from methanol. Phase calculation attempts by using direct methods failed since the data collected with the X-ray source in the laboratory were low resolution due to the presence of many solvent molecules in the structure. Therefore, the X-ray diffraction data sets were measured by synchrotron radiation at four wavelengths, and the phase was calculated by using the multiwavelength anomalous diffraction (MAD) method, [17] by employing the anomalous scattering effect of the bromine atoms. As expected, the accurate phase was successfully determined and the crystal structure of 1 was obtained (Figure 1).In an asymmetric unit of the crystal of 1, four conformationally different molecules were found to exist. In all four, the calixarene moiety maintained a flattened-cone conformation and ...

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A Template‐Assembled Synthetic Protein Surface Mimetic of the von Willebrand Factor A1 domain Inhibits Botrocetin‐Induced Platelet Aggregation

Cited by 16 publications

References 32 publications

Optimal Attachment Position and Linker Length Promote Native‐like Character of Cavitand‐Based Template‐Assembled Synthetic Proteins (TASPs)

Optimal Attachment Position and Linker Length Promote Native‐like Character of Cavitand‐Based Template‐Assembled Synthetic Proteins (TASPs)

Synthetic Peptide Templates for Molecular Recognition: Recent Advances and Applications

Calix[6]arene‐Based Template for X‐ray Crystallographic Analysis of Template‐Assembled Synthetic Proteins

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