Biochemical and environmental sensing with cavitands

Abstract

Post-translational modifications (PTMs) such as histone lysine and arginine residues methylation have a large influence on gene expression being a fundamental part of the histone code. Synthetic supramolecular receptors able to selectively recognized different degree of methylation and isomeric modification states such as asymmetric dimethylarginine and symmetric dimethylarginine for histone code analyses are in highly demand. In this thesis the recognition properties of a Tetraphosphonate cavitands (Tiiii) towards mono and di-methylated arginine were explored via NMR and ITC studies. Moreover, the ability of the Tiiii cavitand in sequestering monomethylated amines exploiting the multivalency approach was investigated. In particular, Tiiii grafted onto ferromagnetic nanoparticles was used to selectively obtain mono-methylated lysine residues onto proteins during the protein methylation reaction. Finally, the Tiiii cavitand was grafted onto gold nanoparticles and its molecular recognition properties towards sarcosine and N-phenethylamine was tested via NOE pumping and STD experiments. Furthermore, an arrayed suite of synthetic hosts and dyes capable of fluorescence detection of oligonucleotide secondary structures was developed using also the Tiiii cavitand. By using cationic dyes that show affinity for both DNA G-quadruplexes and the synthetic hosts, multiple recognition mechanisms can be exploited to create a unique sensing fingerprint, consisting of different fluorescence enhancements in the presence of select DNA strands. Multivariate analysis of these sensing fingerprints enables discrimination between highly similar G4 structures of identical length and topological type. The excellent selectivity of the array allows an easy differentiation and classification of the G4 structures at the same time, in a simple, non-

invasive manner by number, sequence differences at the 3’ or 5’ also with G4s that display the same folding topology. Cation-driven conformational changes in G4 folding topology can be detected by the arrayed sensor. It is even capable of detecting changes in G4 folding pattern in different types of complex media, including fetal bovine serum, and can selectively detect changing concentrations of G4s in mixtures of multiple nucleotides. Amino acids (AAs) represent an ideal playground for testing complexation ability and selectivity of synthetic receptors, due to their biological relevance and chemical diversity. The recognition of aromatic AAs was explored by using water soluble tetraquinoxaline cavitands (QxCav) and benzopyrazine cavitands (BzPyCav). 1H NMR experiments, ITC analyses and solid state studies were performed in water to probe the ability of these cavitands in recognizing L-Phenylalanine and L-Tryptophan. The capability of the cavitand receptors to recognize aromatic guests was explored as well at the solid/liquid interphase by grafting QxCav onto the surface of gold nanoparticles. Finally, environmental sensing field was investigated since highly selective carcinogenic benzene detection is socially relevant and technologically challenging, due to the concurring requirements of high selectivity and extreme sensitivity. Recently, we have developed a MEMs based device in which the specifically designed EtQxBox cavitand acts at the same time as highly sensitive preconcentrator for BTEX and GC-like separation phase, allowing for the selective desorption of benzene over TEX by applying a smart temperature program on the EtQxBox mesh. EtQxBox, obtained by covalently linking together the four quinoxaline walls, strongly retains BTEX to the point that BTEX can be exhaustively desorbed only at temperatures higher than 250 °C, thus reducing the lifetime of the device. Along this line we synthesized a new quinoxaline-based cavitand, where only two quinoxaline walls are linked together to reduce the binding strength of our receptor towards BTEX, allowing their decomplexation at lower temperatures. Similarly, synthetic studies for the development of quinoxaline-based deep cavitands were performed with the intent of maximizing the affinity of the receptor towards the sole benzene.