Metal-based chemosensors for amino acids, peptides, and nucleotides

An organometallic 4d transition metal complex [CpRhCl2]2, together with commercially available dyes, was used to construct indicator displacement assays (IDAs) for the detection of peptides, amino acids, and nucleotides. The combination of the CpRh complex with the dye azophloxine was found to form a chemosensing ensemble for the sequence-selective detection of histidine- and methionine-containing peptides in water at neutral pH. A strong interaction of the rhodium complex with peptides bearing histidine or methionine residue in the position 1 or 2 allowed to detect these peptides down to a concentration of 0.3 µM. The same organometallic complex and three commercially available dyes were employed to compose an array of IDA chemosensors for the detection of natural amino acids. We found that the variation of the pH can effectively be used to modulate the selectivity of an IDA sensor. An excellent discrimination of 20 amino acids was achieved. The combination of [Cp*RhCl2]2 with the dyes gallocyanine, evans blue, and mordant yellow 10 forms a multicomponent indicator displacement assay (MIDA), which can be used to sense low millimolar concentrations of nucleotides and the pyrophosphate anion in buffered aqueous solution. Moreover, the MIDA allows to simultaneously determine the concentrations of adenosine triphosphate and pyrophosphate/cyclic adenosine monophosphate with a single UV-Vis measurement. In the second part of our work we investigated whether dynamic combinatorial libraries (DCLs) can be used for sensing purposes. We found that the dyes arsenazo I, methylcalcein blue, and glycine cresol red, in combination with the metal salts CuCl2 and NiCl2, form a DCL of differently colored metal-dye complexes. The addition of an analyte able to interact with a member(s) of such a library, results in a re-equilibraion easily detectable by UV-Vis spectroscopy. Using dipeptides as analytes we demonstrated that this way of sensing is very effective – even closely related peptides such as regio- (Ala-Phe vs. Phe-Ala) and stereo- (Phe-Ala vs. D-Phe-Ala) isomers can easily be discriminated. Additionally, we found that the identity of the best sensor depends on the problem to be addressed.

Chemosensors Based on Metal-Dye Complexes for Bioanalytes

Friederike Gundula Marie Zaubitzer

Multicomponent chemosensors were prepared by mixing the rhodium(III) complex [(η5-C5Me5)RhCl 2]2, nickel(II) chloride and/or copper(II) chloride with commercially available dyes in buffered aqueous solution. The sensors were used to detect various biomolecules, which interact with the components of the sensing ensemble and alter the composition, and hence the color of the sample. The sensors were analyzed by UV/Vis spectroscopy. A mixture of [(η5-C5Me5)RhCl 2]2, three dyes and pyrophosphate was employed to determine in retrospect the time at which the target molecules ADP and ATP were added to the sensing ensemble. An artificial neural network was used to correlate absorbance data with analyte addition times. Dynamic combinatorial libraries of metal-dye complexes prepared from NiCl2, CuCl2 and three dyes were used to distinguish between mixtures of the peptide hormones angiotensin I and angiotensin II. The discriminative power of the sensor was increased by analyzing samples in a time-resolved fashion. Nine primary amines were identified in a sensing ensemble, where amines were first combined with pyridine-2-aldehyde in the presence of NiCl2 or CuCl2, in order to obtain metal-stabilized imine mixtures. A colorimetric output was generated by adding a dye. In an approach to sense conventional sugars in an indicator displacement assay, sugars were transformed in situ into sugar imines prior to the sensing experiment.

EPFL2010

Solution behavior of peptide and peptide-hybrid materials

Harald Nuhn

Peptide based and peptide hybrid materials represent two interesting and challenging classes of biomaterials. Both classes consist of and contain, respectively, one or more polypeptide blocks, endowing the resulting polymers with enhanced self-assembly properties, biocompatibility or novel characteristics. Peptide based materials, solely comprised of amino acids, benefit from their advanced self-assembly properties. In the case of peptide hybrid materials, being conjugates of peptides and synthetic polymers, either the polypeptide or the synthetic polymer block can drive the self-assembly process. Furthermore, the latter class benefits from combining the advantages of peptides and synthetic polymers, thus overcoming limitations related to the use of the individual components. The application of these materials is still challenging because it is not possible to predict their final self-assembly behavior based on their design. In order to facilitate a target-oriented design of novel materials for application such as drug delivery, tissue engineering, or self-healing materials, a detailed understanding of the complex interactions within and between these molecules is essential. The work presented in this thesis addresses these fundamental questions. Over the last decades, a plethora of peptide and peptide hybrid materials has been reported. Chapter 1 gives an overview of selected articles describing different synthesis routes for peptide as well as peptide hybrid materials, showing examples of molecules and their self-assembly into superstructures, and highlighting recent applications. Chapter 2 describes the synthesis and characterization of different peptide hybrid diblock oligomers composed of two classes of peptide sequences covalently attached to a poly(ethylene glycol) (PEG) block. The first class of peptide sequences contains the intrinsic propensity to form coiled coils, whereas the second class is composed of "switch" peptides and is able to adopt both amphiphilic α-helical and β-strand conformations. Upon dissolution, the first class yielded superstructures comprised of dimeric and tetrameric coiled coil aggregates, whereas the second class formed fibrillar assemblies. In both cases, the self-assembly properties were driven by the peptide block, and the final superstructure consisted of a peptidic core wrapped by the synthetic polymer block. Chapter 3 presents a detailed study on the aqueous solution self-assembly of two poly(styrene)n-b-poly(L-lysine)m peptide hybrid oligomers. The aim was to clarify whether the observed rod-like micelles, obtained by direct dissolution of the diblock oligomers in aqueous solution, reflect intrinsic self-assembly properties of the hybrid diblock oligomers or whether they present structures that are energetically trapped due to the glassy nature of the polystyrene block. To address this question, the influence of a variety of sample preparation techniques, including the use of organic solvents, dialysis from an organic solvent and several thermal treatments, was investigated. All treatments resulted in solution aggregates that differed in size and shape from samples prepared by direct dissolution in aqueous solution. Thus, it seems likely that direct dissolution of polystyrene-b-poly(L-lysine) diblock oligomers in aqueous solution results in kinetically trapped, rod-like aggregates, which can be transformed into non-spherical micelles by the appropriate treatment. Chapter 4 contains a comprehensive study on peptide-based materials, investigating four different series of short elastin-like peptides (ELPs) based on the GVGVP motif. The objective was to test whether and how defined changes in the primary structure of short ELPs affect the secondary structure and the lower critical solution temperature (LCST) behavior. Therefore, the number of repeats was altered, and within one peptide of constant length, all valines were successively substituted by the more hydrophobic amino acids isoleucine, leucine and phenylalanine, respectively. In parallel, a theoretical approach based on the partition coefficient log P was used to predict the shift of the LCST as function of chain length and composition. For short ELPs, the LCST is strongly affected by changes in the primary structure. This has also been predicted by calculating log P. An increase in the hydrophobicity resulted in a shift of the LCST towards lower temperatures. Furthermore, it was shown for the first time that the sensitivity of short ELPs towards external factors, such as salt concentration, is determined by the intrinsic propensity of the incorporated amino acids to form either α-helices or β-strands. In summary, the obtained results on peptide based and peptide-hybrid materials provides deeper insights into their self-assembly characteristics and highlights the potential of these materials to be used for technical and medical applications.

EPFL2008

Sensing with Dynamic Combinatorial Libraries of Metal-Dye Complexes: The Advantage of Time-Resolved Measurements

Kay Severin, Friederike Gundula Marie Zaubitzer

A dynamic combinatorial library of metal–dye complexes was obtained by reacting aqueous solutions of the dyes Methyl Calcein Blue, Arsenazo I, and Xylenol Orange with CuCl2 and NiCl2. The mixture gave a characteristic UV-Vis response upon addition of the peptide hormones angiotensin I and angiotensin II. This allowed distinguishing pure samples of peptide hormones from mixtures. The discriminatory power of the sensor was enhanced when the several UV/Vis measurements were performed during the equilibration process of the library.

2009

Solution behavior of peptide and peptide-hybrid materials

Harald Nuhn

EPFL2008