Gabriele Tuchscherer

Template assembled synthetic proteins : condensation of a multifunctional peptide to a topological template via chemoselective ligation

Abstract A chemoselective ligation via oxime bond formation is used for the chemical synthesis of template assembled peptides according to the TASP (Template Assembled Synthetic Proteins) approach. Aminooxyacetylation of the multifunctional partial sequence Lys- Arg- Asp- Ser of lactoferrin and subsequent condensation in aqueous solution with a topological template containing four selectively addressable aldehyde functions as attachment sites gives readily access to the TASP molecule.

Protein design: On the threshold of functional properties

The ultimate goal in protein de novo design is the creation of novel macromolecules with tailor-made receptor, sensory, and catalytic functions. Despite considerable progress in understanding basic rules of secondary structure formation and protein stability, the well-known protein folding problem is still far from being solved and, in general, only a limited number of designed proteins are folded uniquely. In this article the state-of-the-art in protein design is demonstrated on some selected examples, indicating that the construction of protein-like macromolecules mimicking some essential features of natural proteins seems to be within reach. Thus, protein design and mimicry has become an interdisciplinary challenge with most intriguing perspectives.

Synthesis of antiparallel 4α-helix bundle TASP by chemoselective ligation

The use of chemoselective ligation methods and orthogonal protection techniques allows access to Template-Assembled Synthetic Protein (TASP) molecules exhibiting a large variety of packing topologies. This is demonstrated for the synthesis of an antiparallel 4α-helical bundle TASP by condensing amphiphilic peptide blocks, containing aldehyde functions at the C- or N-terminus, to a selectively addressable topological template via oxime bond formation. The resulting antiparallel 4α-helix TASP is obtained in high yield and shows a template-induced helical conformation.

Synthesis of biotinylated glycosulfopeptides by chemoselective ligation

Tyrosine sulfation and O-glycosylation are two post-translational modifications playing an essential role in modulation of biol. activity, protein folding and cellular communication. Here, a novel chem. approach for the synthesis of biotinylated glycosulfopeptides is described based on a combination of acid labile protecting groups, highly acid sensitive resins and two orthogonal chemoselective ligation reactions. [on SciFinder (R)]

Surface grafting onto template-assembled synthetic protein scaffolds in molecular recognition

Creating functional biological molecules de novo requires a detailed understanding of the intimate relationship between primary sequence, folding mechanism, and packing topology, and remains up to now a most challenging goal in protein design and mimicry. As a consequence, the use of well-defined robust macromolecules as scaffolds for the introduction of function by grafting surface residues has become a major objective in protein engineering and de novo design. In this article, the concept of scaffolds is demonstrated on some selected examples, illustrating that novel types of functional molecules can be generated. Reengineered proteins and, most notably, de novo designed peptide scaffolds exhibiting molecular function, are ideal tools for structure-function studies and as leads in drug design.

Engineering of zinc finger and MHC motifs to locked-in tertiary folds

The assembly of helical and β-sheet peptide blocks containing reactive chain ends results inhighly branched chain architectures (‘locked-in folds’) mimicking native tertiary structures.This molecular kit strategy allows to bypass the protein folding problem in protein de novodesign and gives access to protein mimetics of high thermodynamic stability. The validity ofthis concept is exemplified for the design and synthesis of locked-in folds mimicking the zincfinger and MHC folding motifs.

Template assembled synthetic peptides (TASP) as receptor mimetics and ‘locked-in’ tertiary folds

A symposium report. A new concept for the construction of protein mimetics based on the sepn. of functional and structural domains in proteins has been evaluated by the successful design and total synthesis of two prototype TASP mols. [on SciFinder (R)]

Design and synthesis of a chimeric TASP molecule as potential inhibitor in cell adhesion processes

The construction of protein-like folding motifs as structurally stable scaffolds for the introduction of function represents a major goal in protein design. The use of topol. templates allows the bypass of the well-known folding problem of linear polypeptides and offers a way to mimic native packing topologies by the template directed self-assembly of helical and/or b-sheeted peptide blocks. In conceptually sepg. structure from function, a chimeric 4-helix bundle TASP (Template Assembled Synthetic Protein) derived from the ROP protein and the cell adhesion glycoprotein E-selectin has been designed and synthesized, aimed at inhibiting an early stage in cell adhesion processes, in particular leukocyte adhesion. [on SciFinder (R)]

Pseudoproline libraries for tuning inhibitors of SH3 domain mediated protein-protein interactions

Specific protein-protein interactions are essential facets in cellular communication and the formation and specific assembly of multicomponent protein complexes often is regulated by binding to proline (Pro)-rich peptide sequences. Pro-rich ligands adopt a left-handed polyproline II helical conformation (PPII, all trans amide bonds) and bind to a highly conserved patch of aromatic amino acids of e.g. Src homology (SH3) domains [1]. The essential feature of SH3 binding ligands is the consensus sequence Pro-Xaa-Xaa-Pro (Xaa representing variable amino acids). In the search for novel inhibitors, recently introduced pseudoprolines (ΨPro), i.e. Ser, Thr, Cys derived proline-ring structures with enhanced inherent properties of L-Pro, were used to study ligand receptor interactions of Pro-rich peptides [2]. Binding affinities in the order typically found for SH3-mediated interactions and most notably, enhanced binding specificity as well as inhibition of Grb2 SH3 (N)-SoS complex formation (Figure 1) illustrate that ΨPro building blocks exert a dual functionality, i.e. i) increase and optimization of van der Waals contacts and hydrogen bonding to the receptor molecule, and ii) enhancement of the relevant PPII conformation [3]. To further optimize ligand-receptor interactions in the search of potent SH3 ligands, 2C-substituted ΨPro libraries applying post-insertion strategies have been generated allowing for rapid screening of ligands that optimally complement the SH3 topography.

Non native architectures in protein design : Four helix bundles as locked-in tertiary fold

The authors report here prepn. of the sandwiched 4-helix bundle by sequential oxime bond formation. [on SciFinder (R)]