Disulfide-based dynamic combinatorial libraries of macrocyclic pseudopeptides as bio-inspired complex chemical systems

Abstract

Dynamic combinatorial chemistry proposes the creation of a library of compounds (dynamic combinatorial library, DCL) inter-connected through reversible chemical processes. The concentrations in a DCL are determined by differences in the free energy between the constituents and, therefore, the variations in the composition of the library contain valuable information about changes in the stability of the members. The adaptive nature of DCLs can be intimately correlated with a very simple expression of molecular evolution. The main objective of this thesis is to use DCLs as bio-inspired complex chemical systems for the minimalistic experimental modeling of different processes of biological interest.

In Chapter 1, fourteen new pseudopeptidic dithiols were designed, synthesized and fully characterized in order to be used as bipodal building blocks (BBs). Their design is based on a C2-symmetric scaffold consisting of a central m-phenylenediamine chromophore that rigidly joins two identical arms, each formed by an amino acid with a mercaptoacetyl moiety attached to the N-terminus. The pseudopeptidic nature provides the BBs with peptide-like information, differently charged functional groups and chiral information. In Chapter 2, suitable experimental conditions were developed for the generation of disulfide-based DCLs from the mixture of the synthesized BBs. For this purpose, the use of DMSO as a co-solvent has beneficial effects in the thiol-disulfide dynamic covalent chemistry. Apart from the general gain in the aqueous solubility of the organic molecules, it promotes the thiol oxidation, highly reducing the reaction time for the disulfide formation. Besides, it accelerates the disulfide exchange, allowing the system to fully equilibrate even at slightly acidic pH. In Chapter 3, a minimalistic DCL was used to reproduce adaptive trends described for the evolution of biological systems. Thus, the addition of salt to a dynamic library of macrocyclic pseudopeptides induces the amplification of those species concentrating anionic amino acids, with the Asp derivatives showing a better salt-adaptation than the Glu counterparts. Structural studies suggest a folded conformation for the amplified members and reveal the selection of those species showing a smaller accessible surface area. The adaptive process is driven by the increase of the ionic strength and has a remarkable resemblance with the natural evolution of the proteins of halophilic microorganisms for surviving in hypersaline media. In Chapter 4, the same external stimulus was studied in a larger DCL consisting of 21 differently charged dimeric macrocycles. The salt-induced adaptation of this complex system was characterized in a top-down fashion by the dynamic deconvolution into the minimal components.

Additionally, structural studies were performed for selected species. The salt-response of the members of the library can be classified in different families attending to the charges, and the behavior of each member is determined by a combination of its structural information and the co-adaptive relationships with the other members of the complex network.

Finally, in Chapter 5, a simple DCL consisting of homo- and heterochiral dimeric pseudopeptides was used to study how the chiral information is transmitted from the molecular to the macromolecular level. A decrease in the polarity of the medium induces a homochiral self-sorting process driven by polar intramolecular interactions. Additionally, the homochiral selectivity also increases with the temperature, indicating a positive entropic contribution. Preliminary NMR experiments suggest significantly different conformations for the homo- and heterochiral species.

Overall, the adaptive nature of DCLs, together with a suitable bio-inspired design, have demonstrated to allow the minimalistic experimental modeling of different processes of biological interest such as the natural evolution of the halophilic proteins, the co-adaptive relationships in a complex network and the homochiral self-sorting phenomenon.

Les quimioteques dinàmiques (DCLs de “dynamic combinatorial libraries”) estan formades per una mescla de compostos interconnectats per mitjà de processos químics reversibles. Aquests sistemes dinàmics presenten la capacitat d’adaptar la seva composició a la presència d’un estímul extern. L’objectiu principal d’aquesta tesi és utilitzar DCLs com a sistemes químics complexos pel modelatge experimental de diversos processos d’interès biològic.

En el Capítol 1 es van dissenyar, sintetitzar i caracteritzar catorze ditiols amb informació estructural de tipus peptídic, cadenes laterals amb diferents càrregues i informació quiral. A continuació, en el Capítol 2, es van establir unes condicions experimentals adequades per a generar DCLs de disulfurs a partir de l’oxidació dels ditiols sintetitzats. En aquest sentit, l’ús de DMSO com a codissolvent orgànic va demostrar tenir una sèrie d’efectes beneficiosos. Seguidament, en el Capítol 3, es va utilitzar una DCL minimalista de disseny bioinspirat per a reproduir tendències adaptatives pròpies de processos evolutius biològics. Així, es va observar que els canvis que provoca l’increment de la salinitat en la composició d’una DCL de pseudopèptids macrocíclics, tenen una notable similitud amb l’evolució natural de les proteïnes dels microorganismes halòfils. En el Capítol 4 es va estudiar l’efecte del mateix estímul extern en la composició d’una DCL complexa formada per espècies amb diferents càrregues. S’evidencià que el comportament de cada membre de la quimioteca està determinat per la seva informació estructural i per les múltiples relacions coadaptatives que aquest estableix amb la resta de membres de la xarxa molecular. Finalment, en el Capítol 5, es va utilitzar una DCL minimalista formada per espècies estereoisomèriques per estudiar el fenomen d’autoordenació homoquiral.