1,685 research outputs found
Synthesis of cyclic peptides and peptidomimetics by metathesis reactions
Full text linkThis review surveys developments in the field of ring-closing metathesis and cross-metathesis reactions applied to the synthesis of constrained amino acids, peptides, and peptidomimetics. Examples, in which metathesis is used as one of the synthetic tools to arrive at the desired peptide molecules as well as examples that describe in-depth optimized metathesis protocols, will be discussed. Currently, metathesis reactions are well-accepted synthetic tools within the field of peptide chemistry and provide peptides unprecedented properties like conformational, metabolic, and chemical stability and improved bioactivity
Towards Bio-inspired and Functionalized Peptide Materials
No full textPeptide-based materials constitute a class of molecules that play an important role in many biological processes and are utilized by many organisms to interact with their environment. One of the most well-known examples is spider silk, a material produced by web-spinning spiders composed of repeating stretches of amino acids that has a very high tensile strength. Many peptide-based materials are composed of repeating stretches of certain amino acids that confer function and (macroscopic) structure to these materials. In the first three research chapters a peptide material based on naturally occurring antifreeze molecules from arctic fish is described. In nature this material is composed of repeating stretches of a glycosylated tripeptide. Our mimic is synthesized by a copper-catalyzed polymerization reaction from a modified glycotripeptide with polymerization handles. This polymerization reaction introduces a non-natural element in the peptide backbone that was known to mimic the natural amide bond in previously published examples to a certain extent. A one-pot reduction and polymerization yielded a mixture of oligomers that could be separated by size. Upon analysis of the antifreeze activity of the resulting antifreeze mimics by an ice recrystallization experiment, the polymeric material was shown to have a strongly reduced antifreeze activity compared to the natural compound and a synthetic derivative synthesized previously. Structural analysis by circular dichroism showed a similar structure to the previously described and potent synthetic antifreeze glycopeptide, indicating that a small structural change can abolish the potent antifreeze properties of such molecules. The chemistry for introducing a urea moiety that can be incorporated into the antifreeze glycopeptides to enhance the interaction with the ice-lattice is described, using a 4-chlorophenyl glycoside synthon that can react with amines under mild conditions to form a ureido glycoside linkage. As a case study, this chemistry was used to synthesize a urea-containing glycopolymer based on the previously synthesized mimic. In chapter five a modification of the naturally occurring antibiotic nisin is described, wherein commercially available nisin was conjugated on the C-terminus with propargylamine, a handle for copper catalyzed click chemistry. In a second step this modified nisin could be coupled to fluorescent reporter molecules, and as a proof-of-concept, a small molecule with two azide handles. The successful synthesis of the dimeric nisin structure and retention of antimicrobial activity of all conjugates indicated the possibility to easily incorporate nisin in a functionalized (peptide) material or surface to obtain antimicrobial peptide based materials. In chapter six a dicysteine motif is described, which easily oxidizes to a cyclocystine. Initially found in a peptide derived from Tamm-Horsfall protein (F991) that was indicated to bind free-light chains, it became apparent that in serum this peptide rapidly formed the cyclocystine moiety. Attempts to use this structural motif in the beta-sheet forming human islet amyloid polypeptide as an oxidative switch are described. Preliminary results show a distorted structure by circular dichroism, but the resulting peptide still formed similar structures in aqueous buffer. Further attempts at increasing the change upon oxidation are described
Protein mimics by molecular scaffolding of peptides
No full textThe research in this thesis aimed at the development of a new route for the synthesis of discontinuous epitope mimics. Mimicry of discontinuous epitopes is often achieved through mounted peptides corresponding to the epitope sequence on a molecular scaffold. In this thesis a new method for the synthesis of these mimics was developed by using sequential introdcution of peptides onto the scaffold using CuAAC chemistry. This resulted in a synthesis method that was convergent, efficient, and easily adaptable. Using this method mimics of epitopes of the Pertussis protein and the gp120 protein were synthesized. The gp120 mimics were able to competitively inhibit the binding of natural gp120 to CD4. Furthermore, the importance of peptide cyclization was investigated. Mimics containing linear peptides were able to inhibit the gp120-CD4 binding but to a somewhat lesser extent. However, the stability in serum of mimics consisting of cyclic peptides was far better than the linear mimics, which shows the benefits of the use of cyclic peptides in protein mimic
Synthesis of Triazole Bridged Vancomycin Mimics
No full textCyclic peptides are an important class of compounds with broad biological activities. Both natural as well as synthetic cyclic peptides have been recognized as a great resource and inspiration for drug discovery. Macrocyclization is recognized as an efficient way to restrict the conformational freedom of a peptide which often leads to increased affinity and selectivity. However, only a limited number of approaches for efficient peptide macrocyclization are presently available. Vancomycin is the most representative member of a family of glycopeptide antibiotics, which are the most important class of drugs for the treatment of resistant bacterial infections. The ongoing development of novel synthetic methodology to access conformationally restricted cyclic peptides encouraged us to develop effective approaches to mimic the bioactive conformation of vancomycin as closely as possible. This thesis describes the application of CuAAC and RuAAC macrocyclization for the synthesis of vancomycin mimics. The synthesis of cyclic 1,4-triazole-bridged tripeptides as vancomycin DE-ring mimics was described. Based on the successful synthesis of a series of linear tripeptides functionalized with alkyne and azide groups, the macrocyclization using CuAAC chemistry was successfully achieved. With the TBTA-promoted CuAAC macrocyclization strategy, two cyclic monomeric tripeptides, derived from ornithine and lysine, were successfully synthesized. As a relevant follow-up study after the successful application of CuAAC for the synthesis of the vancomycin DE-ring mimics, a RuAAC click-type macrocyclization protocol described, and a series of cyclic 1,5-triazole-bridged tripeptides as vancomycin DE-ring mimics was synthesized in good yield. The most constrained cyclic tripeptide obtained had only 13 atoms in the macrocyclic ring, while the lysine-derived tripeptide was of the same size as the vancomycin DE-ring containing 16 atoms. The RuAAC macrocyclization proved to be effective for the synthesis of 1,5-disubstituted triazole-containing cyclic tripeptides, and was superior compared to the CuAAC-based click cyclization with respect to the prevalent synthesis of monomeric cyclic peptides. With the successful application of the RuAAC macrocyclization for the synthesis of the DE-ring mimics of vancomycin, the synthesis of 1,5-triazole-bridged bicyclic vancomycin CDE-ring peptidomimetic using RuAAC macrocyclization was successfully achieved. The synthesis of a linear hexapeptide functionalized with alkyne and azide groups was carefully optimized and the synthesis could be accomplished within 10 steps with good to excellent yield. The RuAAC macrocyclization was successfully developed to synthesize the 1,5-triazole-bridged bicyclic peptidomimetic. The antibacterial activity of the synthesized bicyclic compound was investigated, but no obvious inhibition of the bacterial growth was observed. In general, the investigation as described in this thesis showed the successful application of CuAAC and RuAAC macrocyclization strategy for the synthesis of vancomycin DE- and CDE-ring mimics. Especially, the RuAAC chemistry turned out to be a very efficient strategy for the synthesis of small cyclic peptides with excellent intramolecular selectivity. Although the synthesized peptidomimetics were not biologically active, the methodology developed here could be applied for the structural optimization of cyclic peptides that lead to biologically interesting targets
Semi-synthesis of nisin-based peptide antibiotics
No full textThere is a growing need for novel antibiotics since there are more and more cases of infections caused by resistant bacteria. Possible novel antibiotics are antimicrobial peptides, especially the lantibiotic nisin. Lantibiotics are ribosomally synthesized cationic peptides that contain several unnatural amino acids like dehydroalanine (Dha), dehydrobutyrine (Dhb) and have multiple cyclic structures derived by thio ether bonds or lanthionines, which are very important for its antimicrobial activity. Nisin can inhibit the cell-wall synthesis by binding to lipid II (an important lipid in the bacterial cell-wall) with its N-terminal A and B rings. After binding, nisin is able to form a pore-complex by inserting its C-terminus (C, D/E rings) into the bacterial cell membrane resulting in a collapse of vital ion gradients. Although nisin holds a promising template for novel peptide-based antibiotics, synthesis of nisin-like structures is not trivial and especially the introduction of the lanthionine rings is challenging. Moreover, the lanthionine bridges are oxidation sensitive and the approach as described in this research to replace the lanthionines by dicarba bonds via ring-closing metathesis (RCM) could be used to access stabilized nisin-like structures via (chemical) synthesis. In the first part of this thesis, the design, synthesis and evaluation of nisin fragments is described. Native nisin fragments were prepared by using optimized enzymatic and chemical cleavage methods, and two previously unknown cleavage sizes were discovered. Furthermore, through solution and solid phase peptide synthesis, mimics of nisin fragments were successfully prepared replacing the lanthionines by dicarba bonds using RCM. Native and synthetic nisin fragments were assembled via copper-I catalyzed click chemistry to obtain full-length nisin hybrids and their biological activities were evaluated. This orthogonal ligation strategy via click chemistry was also successfully applied on full-length native nisin to ligate several conjugation partners, to obtain biologically active fluorescent nisin derivatives, which are important tools to further study nisin’s mode of action. Finally, a lipophilic amino acid was designed, synthesized and incorporated into the short antimicrobial peptide anoplin as a model peptide. This strategy demonstrated an approach to improve the antimicrobial activity by increasing the lipophilicity of anoplin, while retaining its selectivity for bacterial membranes, and could potentially be applied as a general strategy to improve the activity of membrane-acting peptides. The synthesized nisin dicarba AB analogs proved to be biologically active and the importance of the native backbone structure was shown. The nisin hybrids were biologically active, however, they lacked pore-formation activity possibly due to a non-optimal C-terminus. The results of this thesis provide valuable information about the importance of each individual ring structure, which gives more insight into the potential of nisin to improve its metabolic stability and thereby its antibiotic properties
Concurrent formation of furan-2,5- and furan-2,4-dicarboxylic acid: unexpected aspects of the Henkel reaction
No full textThe concurrent formation of furan-2,5- and furan-2,4-dicarboxylic acid under solvent free conditions via a disproportionation reaction is described. By reacting potassium-2-furoate at 260 °C in the presence of 22 mol% of (Lewis acidic) catalysts like CdI2 or ZnCl2, potassium-2-furoate is disproportionated to furan and furandicarboxylic acids. Besides furan and furan-2,5-dicarboxylic acid (2,5-FDCA) as the main products, furan-2,4-dicarboxylic acid (2,4-FDCA) is also formed as a by-product. Experimental evidence has been obtained that, under the reaction conditions applied, 2,5-FDCA and 2,4-FDCA are formed by separate reaction pathways. Selectivity towards the different FDCA isomers is affected by the type of catalyst used. Single-crystal X-ray analysis shows that 2,4-FDCA has a more ‘linear’ character compared to 2,5-FDCA and hence is structurally more comparable to terephthalic acid (TA), making it an interesting monomer for synthetic polyesters
Immobilization of stabilized antimicrobial peptides into a bactericidal hydrogel coating
No full textThis thesis describes the design and synthesis of a bactericidal poly(ethylene glycol)-based (PEG) hydrogel coating with covalently attached antimicrobial peptides (AMP) stabilized against proteolytic degradation. As such, mimics of the highly active AMP HHC10 (H-KRWWKWIRW-NH2) were designed for optimal stability in human serum while retaining strong antimicrobial activity against Staphylococcus aureus and Staphylococcus epidermidis, the major causative agents of biomaterial associated infection. In order to investigate the selectivity of the AMPs, their hemolytic activity was determined. A N-terminal cysteine facilitated thiol−ene chemistry for a fast, single-step immobilization/photopolymerization strategy. The antimicrobial activity of the resulting thin layer hydrogel coating on a PET surface was established using the Japanese Industrial Standard (JIS) Z2801 assay, showing complete killing (>99.9%) of inocula of S. aureus, S. epidermidis, and E. coli. The bactericidal hydrogel was molecularly characterized via Coomassie and Lowry assay protein staining agents as well as by X-ray photoelectron spectroscopy. To gain further insight into the biological stability, the hydrogels were incubated with human serum prior to activity testing without loss of activity. These studies revealed a promising bactericidal hydrogel with good stability under physiological conditions. However, the in vivo activity in mice could not be demonstrated and should be further investigated to prove the full potential of this methodology
New rigid spacers for multivalency studies: strong inhibition of Pseudomonas aeruginosa lectin LecA
No full textEspecially in the case of a chelating binding mode, it is necessary to find the optimal linker. The results obtained led to the conclusion that, despite the difficulties encountered during the design of a rigid linker, a fixed geometry could bring an enhancement in the affinity and a higher selectivity. The lectin LecA from Pseudomonas aeruginosa is an extremely interesting target, not only because of its biological relevance but also for its suitability to study the chelating multivalency effect. Furthermore, the affinity for LecA showed to be dependent on the geometry of the multivalent system and it seemed to be an optimal target to investigate the role of a rigid linker. This thesis covers the design and the synthesis of new rigid spacers used to form divalent systems. The spacer designs were based on a triazole-glucose repeating unit or on phenylene-ethynylene repeating unit. The inhibitors obtained were tested with LecA to evaluate a correlation between rigidity and affinity. The first spacer taken into account was based on a triazole-glucose repeating unit and in Chapter 2 the attempts to synthesize a suitable building block were described. The ideal building block required reproducible and high-yielding reactions. The successfully synthesized building block was used in Chapter 3 to synthesize rigid spacers. The strategy was based on the reiteration of three reactions: copper alkyne-azide cycloaddition (CuAAC) and introduction of the azide moiety via a triflate. The spacers synthesized were characterized by a different number of units, ranging from 2 to 4. The galactoside ligands coupled to the spacers exhibited different degrees of freedom. Furthermore, a PEG-based divalent system was synthesized as a flexible reference to compare with the rigid systems. The divalent inhibitors obtained were evaluated with lectin LecA. In Chapter 4 a different spacer design is described. The repeating unit was based, this time, on a phenylene-ethynylene moiety. The syntheses of the building blocks is described. The strategy consisted of alternating Sonogashira reactions and silyl-protecting group removal. The spacers obtained contained two to five phenylene-ethynylene unit. The three-unit spacer was coupled to a galactoside ligand and its potency as a LecA inhibitor was determined. In the last chapter, Chapter 5, a new and faster strategy to build rigid spacers based on the triazole-glucose unit is described. To increase the solubility of the protected spacer a new building block containing a silyl group was synthesized. The three different length spacers, from two to four units, were coupled with galactose ligands with different aglycon chains. Their inhibitiory potencies were tested with LecA and, for some compounds, also the Kd values were determined by ITC in selected cases
Design and synthesis of discontinuous protein binding site mimics of HIV gp120
No full textEssentially all cellular processes are mediated by protein-protein interactions and detailed knowledge about these interactions can aid in determining biological functions or provide opportunities for the treatment of human disease. Extensive research has been performed on the development of modulators of protein-protein interactions, since these molecules could provide better understanding of the mechanisms of molecular recognition, lead to the identification of drug targets or provide access to novel therapeutics. Peptide-based mimics of protein binding sites are promising candidates for this purpose. Despite the progress that has been made in the field of protein mimicry and the increasing understanding of protein-protein interactions, the development of molecules that can modulate these interactions remains a challenging task because of the complex nature of protein binding sites. Protein-protein interaction sites often consist of discontinuous epitopes. An important example of such an interaction is the CD4 binding site of HIV gp120. Since gp120 plays a central role in the ability of HIV to enter cells, it is an interesting target for vaccine research. Therefore, in this thesis the development of a fast and reliable method for the synthesis of collections of discontinuous epitope mimics is described. The method comprises the conjugation of three different cyclic peptides, representing the epitopes, to a scaffold molecule, using the copper(I)-catalyzed alkyne-azide cycloaddition reaction. The main characteristic of our method is the rapid generation in a single experiment of a mixture of multiple discontinuous epitope constructs. Thus, a diversity of combinations of cyclic peptides on a scaffold molecule is accessible in a reproducible manner. These different constructs can be conveniently separated by preparative HPLC and characterized with mass spectrometry. The mixture and the individual constructs were tested for CD4-binding affinity with ELISA for evaluation of their ability to act as gp120 mimics. This synthetic procedure provides rapid access to a diversity sophisticated peptide biomolecular constructs. Moreover, the synthesis and activity screening of multiple products in one approach enhances the probability to rapidly generate a starting point for developing synthetic vaccines derived from gp120 or neutralizing antibodies targeting CD
Het feest van de suikers en andere gasten
No full textKoolhydraten spelen een veel belangrijkere rol in biologische processen dan altijd werd aangenomen. Prominent aanwezig op celoppervlakken, zijn zij betrokken bij veel communicaties en interacties. Soms zijn deze interacties ongewenst en ontstaan ziektes. Het maken van moleculen die hier kunnen helpen is het doel. De ongebruikelijke taal van de suikers vraagt een specifieke synthetische aanpak met aaneengekoppelde suikersystemen. Naast de route naar geneesmiddelen zijn diagnostiek en het oplossen van biologische vraagstukken belangrijke mogelijkheden
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