Kubik Group

Research - Cyclopeptides

Introduction

Cyclopeptides are ubiquitously present in Nature. Their high physiological activity is often a result of the stabilization of certain bio-active conformations upon cyclization while the cyclic structure simultaneously protects against degradation by proteases.

Yet, cyclopeptides cannot only act as (small) substrates for (larger) receptors, the macrocyclic structure also allows them to serve as receptors for (smaller) guest molecules that can be bound inside the cavity. The natural cyclodepsipeptide valinomycin is a prominent example of such a receptor whose antibiotic activity is due to the ability to complex and transport potassium ions along bacterial cell membranes.

Although pioneering work on the development of cyclopeptide derived macrocyclic ligands was carried out in the group of E. Blout already between 1970 and 1990, the concept of using cyclopeptides as synthetic receptors in molecular recognition was not widely accepted in spite of the fact that cyclopeptide-based receptors possess various advantages with respect to macrocyclic receptors derived from, for example, crown ethers, calixarenes, or cyclodextrins. Specifically,

  • cyclopeptides are structurally closely related to natural systems,
  • their ring size is easily variable,
  • they are prepared by sequential synthesis which allows for the introduction of various binding sites along the ring in a defined arrangement,
  • their subunits can be varied in a wide range,
  • and they are chiral.

These considerations motivated us to systematically study the host-guest chemistry of cyclopeptides. In this context, we concentrated on compounds containing natural and non-natural aromatic amino acid subunits in an alternating sequence along the ring with the rigid aromatic subunits mainly serving to reduce conformational flexibility.

Cyclopeptid

We showed that such peptides are efficient synthetic receptors for cations and anions. Receptor affinity can be regulated by controlling the favored conformation in solution. While some peptides interact only with one component of an ion-pair, others are able to complex both ion-pair components simultaneously. The introduction of additional binding sites along the ring furnished host molecules for neutral guests, for example carbohydrates.

Our work thus clearly demonstrates the potential of such cyclopeptides as synthetic receptors and we expect a number of interesting applications for these compounds.

Ring Size

So far, cyclic tetra-, hexa-, and octapeptides containing 3-aminobenzoic acid derived subunits have been synthesized in the group. Since hexapeptides were used in the majority of binding studies, these peptides were structurally varied the most. Acyclic natural α-amino acids must be used for the preparation of cyclic octapeptides. If the natural amino acid is proline, cyclic tetrapeptides are accessible. The crystal structure of one of these tetrapeptides reveals that the amide groups at the proline residues adopt the cis conformation.

Kristallstruktur

[For an interactive version of the crystal structure click here]

The crystal structure also shows that such tetrapeptides lack a well-defined cavity. Cation affinity is therefore lower than that of corresponding hexapeptides. Tetrapeptides are, however, promising candidates for the development of pincer-type receptors.1 In his context, we have developed a bisboronic acid based on a cyclic tetrapeptide that binds glucose in aqueous solution enantioselectively.2

References

  1. S. Pohl, R. Goddard, S. Kubik Tetrahedron Lett. 2001, 42, 7555-7558.
  2. G. Heinrichs, M. Schellenträger, S. Kubik Eur. J. Org. Chem. 2006, 4177-4186.

Cation Recognition

Cyclic hexapeptides with 3-aminobenzoic acid subunits bind quaternary ammonium ions in chloroform. The cation affinity of a cyclopeptide containing glutamic acid residues is, however, small.1 One reason is the conformational flexibility of this compound that is caused by relatively free rotations around the bonds at the glutamic acid C(α) atoms. As a consequence of these rotations, several dish-shaped cyclopeptide conformations differing in the orientation of the aromatic subunits equilibrate in solution. The peptide is therefore poorly preorganized for cation binding.

conformations

Rotations around one bond at the C(α) atoms is prevented in a cyclopeptide containing proline instead of glutamic acid subunits. In spite of the overall reduced conformational flexibility of this compound rotations are possible at the secondary amide groups because intramolecular hydrogen bonds prevent this motions to occur. The proline containing peptide is still better preorganization for guest binding and forms more stable complexes with quaternary ammonium ions than the glutamic acid containing one.2

conformations

Rotation around the secondary amide groups is prevented in proline-derived cyclopeptides containing substituents in the 4-position of the aromatic subunits, which can form intramolecular hydrogen bonds to the neighboring NH groups. Such peptides thus adopt conformations in solution optimal for complex formation even in the absence of cationic substrates.3

conformations

The highest cation affinity was observed for a peptide with methoxycarbonyl substituents whose crystal structure is depicted below.3,4

crystal

[For an interactive version of the crystal structure click here]

The chirality of the cyclopeptides also allows for enantioselective recognition of chiral quaternary ammonium ions.5

References

  1. S. Kubik J. Am. Chem. Soc. 1999, 121, 5846-5855.
  2. S. Kubik, R. Goddard J. Org. Chem. 1999, 64, 9475-9486.
  3. S. Kubik, R. Goddard Eur. J. Org. Chem. 2001, 311-322.
  4. S. Kubik, R. Goddard Chem. Commun. 2000, 633-634.
  5. G. Heinrichs, L. Vial, J. Lacour, S. Kubik Chem. Commun. 2003, 1252-1253.

Anion Recognition

Anions can bind to the NH groups of cyclopeptides by hydrogen bond formation. Cyclic hexapeptides with 3-aminobenzoic acid subunits and glutamic acid or proline residues adopt conformations with converging NH groups upon complexation of sulfonate that are optimal for anion binding. Hence, the optimal preorganization for anion-binding of a cyclopeptide that prefers such a conformation also in the absence of the guest should cause high affinity. The characterization of the binding properties of a cyclic hexapeptide with L-proline und 6-aminopicolinic acid subunits confirmed this assumption.1

structure

The converging arrangement of the three NH groups of this cyclopeptide is clearly visible in the crystal structure.

crystal

[For an interactive version of the crystal structure click here]

This peptide binds anions such as iodide or phenylsulfonate in DMSO in the form of 1:1 complexes.2 Interestingly, complexation of halides or sulfate is even possible in highly competitive aqueous solvent mixtures, for example in 80% water/methanol. Under these conditions, sandwich-type 2:1 complexes are, however, formed in which the anion is positioned between two interdigitating cyclopeptide rings binding to six NH groups simultaneously. This arrangement was also detected in the crystalline iodide complex. This cyclopeptide represents one of the few neutral systems that allows the complexation of anions in aqueous media.

crystal

[For an interactive version of the crystal structure click here]

Formation of the 2:1 complex in aqueous solvent mixtures is a cooperative process since the formation of the 2:1 complex from the 1:1 complex is associated with a significantly larger equilibrium constant than the formation of the 1:1 complex from the components.3

The 2:1 complexes could be converted into 1:1 complexes by covalently linking two cyclopeptide rings together which improved complexation efficiency significantly.4

structure

Water-soluble bis(cyclopeptides) were synthesized recently, which bind anions in water.5 Their anion affinity in water and organic solvents could be correlated with the properties of the anions and the solvent.

References

  1. S. Kubik, R. Goddard, R. Kirchner, D. Nolting, J. Seidel Angew. Chem. 2001, 113, 2722-2725; Angew. Chem. Int. Ed. Engl. 2001, 40, 2648-2651.
  2. S. Kubik, R. Goddard, S. Otto, S. Pohl, C. Reyheller, S. Stüwe Biosens. Bioelectr. 2005, 20, 2374-2375.
  3. S. Kubik, R. Goddard Proc. Natl. Acad. Sci. U.S.A. 2002, 99, 5127-5132.
  4. S. Kubik, R. Kirchner, D. Nolting, J. Seidel J. Am. Chem. Soc. 2002, 124, 12752-12760.
  5. F. Sommer, Y. Marcus, S. Kubik ACS Omega 2017, 2, 3669-3680.

Ion-Pair Recognition

While cation affinity of the cyclopeptides is due to the aromatic subunits and the carbonyl groups along the macrocyclic cavity, anions can bind to the cyclopeptide NH groups. Some cyclopeptides bind both components of an ion-pair simultaneously. A glutamic acid containing cyclic hexapeptide, for example, interacts with sulfonates or phosphonates by adopting a conformation with all six NH groups converging toward the center of the macrocyclic cavity. Complexation of the anions increases cation affinity because it improves preorganization of the peptide for cation binding and allows the included cation inside the cyclopeptide cavity to interact with the amino acid residues and the anion.1

Anionenkomplexe

A similar behavior was observed for the proline-containing cyclopeptide. In this case, however, even weakly coordinating anions can be bound because, in contract to the glutamic acid containing derivative, the NH groups of this peptide are not involved in intramolecular hydrogen bonds in the absence of guests.2 The simultaneous interaction with both components of the ion pair is clearly visible in the crystal structure of the N-methylquinuclidinium iodide complex.

Kristallstruktur

[For an interactive version of the crystal structure click here]

Yet, a specific complexation of the anion is not required for an anion effect on cation complexation. Also anions that are not bound by the cyclopeptide but are closely associated to the cation in organic solvents can influence cation affinity. Investigations carried out in collaboration with the group of Jerôme Lacour showed, for example, that the stability of the complexes of a cyclopeptide with a chiral quaternary ammonium ion does not only depend on the absolute configuration of the cation but also on that of the chiral TRISPHAT counterion, which does not specifically bind to the cyclopeptide.3

References

  1. S. Kubik J. Am. Chem. Soc. 1999, 121, 5846-5855.
  2. S. Kubik, R. Goddard J. Org. Chem. 1999, 64, 9475-9486.
  3. G. Heinrichs, S. Kubik, J. Lacour, L. Vial J. Org. Chem. 2005, 70, 4498-4501.

Carbohydrate Recognition

We are also interested in the development of receptors for neutral substrates on the basis of cyclopeptides. The cavity dimension of a hexapeptide with L-proline and 3-aminobenzoic acid subunits is, for example, well suited for the inclusion of a monosaccharide. This peptide lacks functional groups along the cavity, however, with which such a substrate can interact.

We have therefore prepared cyclopeptides derivatives with additional binding sites in the 5-position of the aromatic subunits. The corresponding substituents were chosen to mimic the binding motifs used in natural systems for the recognition of carbohydrates. They thus contain free carboxylate groups that can form hydrogen bonds to the hydroxyl groups of sugars.

carbohydrate

All cyclopeptides synthesized form 1:1 complexes with various monosaccharides in 4% methanol/chloroform. The observed complex stabilities are acceptable if one considers that methanol molecules, present in a ca. 10.000 fold excess in the solvent mixture used, compete with the sugars for the receptor binding sites. Binding selectivity of the cyclopeptides with respect to certain sugar anomers or epimers is low, however.1 Interestingly, the same cyclopeptides bind protected arginine derivatives even in water.2

References

  1. J. Bitta, S. Kubik Org. Lett. 2001, 3, 2637-2640.
  2. J. Bitta, S. Kubik J. Supramol. Chem. 2003, 1, 293-297.

Dynamic Combinatorial Chemistry

We have used dynamic combinatorial chemistry in close collaboration with the group of Sijbren Otto for the optimization of the anion affinity of covalently linked bis(cyclopeptides). In this context, a cyclopeptide disulfide was equilibrated under suitable conditions with different dithiols. Addition of anionic templates, for example sulfate (as potassium sulfate), caused the amplification of receptors in this dynamic library that possess high affinity for this template.1

dynamic

Two of the amplified receptors were synthesized and their affinity toward iodide and sulfate was compared with that of a bis(cyclopeptide) in which the receptor subunits were linked via adipic acid. Both bis(cyclopeptides) bound the two anions ca. one order of magnitude more strongly than the adipic acid containing derivative.

The cooperative action in guest binding of the two cyclopeptide rings is clearly visible in the crystal structure of the sulfate complex of one receptor identified by dynamic combinatorial chemistry.2

crystal

[For an interactive version of the crystal structure click here]

Doubly-linked bis(cycopeptides) that are better preorganized for anion binding in comparison to the corresponding singly-linked derivatives were shown to possess anion affinities in aqueous solvent mixtures (acetonitrile/water 2:1) approaching the nanomolar range.3

References

  1. S. Otto, S. Kubik J. Am. Chem. Soc. 2003, 125, 7804-7805.
  2. Z. Rodriguez-Docampo, S. I. Pascu, S. Kubik, S. Otto J. Am. Chem. Soc. 2006, 128, 11206-11210.
  3. Z. Rodriguez-Docampo, E. Eugenieva-Ilieva, C. Reyheller, A. Belenguer, S. Kubik, S. Otto Chem. Commun. 2011, 47, 9798-9800.

Last change: 21-04-14. Email

Top