Porous peptide frameworks generated by stacking non-self-complementary β-sheets

"One of major approaches in the design of cavity space in the solids utilizes non-self-complementary molecules[1]. The irregular shape of the molecules and/or specific directionality of potential H-bonds prevent close packing of the molecules and yields various architectures hosting a second component, from inclusion compounds and co-crystals to complex non-crystalline patterns in biology. The strategy of non-self-complementary molecules has been extended in our studies to 2D supramolecular polymers based on short peptides[2]. The formation of the peptide layer with a desired overall geometry is controlled by strong, charge-assisted H-bonds (arrows in the Figure) in a β-sheet-like network as well as the segregation of hydrophobic amino acid residues into the interlayer space. The H-bonds add stability to the whole architecture while the hydrophobic groups keep the stacking layers at a distance that generates a cavity space available to a second component (encircled ""G"" in the Figure). A wide range of inclusions and co-crystals have been prepared in our group based on a series of dipeptides and higher peptide oligomers. For example, the incorporation of various organic solvents and bioactive molecules have been demonstrated for leucyl-alanine and similar dipeptides: alcohols, amides, phenols, pyridines, polyols, vitamins, scents and flavors. The crystal structure studies reveal a surprisingly persistent structural motif that can be used for engineering of crystalline materials with a specific property. We believe this type of peptide matrix may be utilized in the solid state organic synthesis [3] as reactive molecules of the second component can be oriented in a predictable way with respect to each other. "

Solid state intramolecular cyclization of leucyl-alanine and alanyl-leucine

Solid state organic synthesis is a future alternative to traditional, solution-based laboratory and industrial synthetic procedures. Solvent-free synthetic methods allow for quantitative yields, high stereospecificity, need no solvent, and are easy to conduct. They may contribute to green economy by reducing pollution, cutting the consumption of energy, and lowering the cost of production of various organic compounds. Although solid state reactions have been reported for almost all main classes of organic compounds and reaction types[1], the reactivity of peptides in the solid state has not been well explored. One potential product of the solid state transformation of dipeptides is 2,5-diketopiperazines (DKPs), the cyclic forms of dipeptides. They have attracted attention due to their high biological activity and use in medicinal chemistry[2,3]. In this study, we investigated the thermally induced intramolecular cyclization of leucyl-alanine and alanyl-leucine in the solid state. The reaction was conducted in a range of experimental conditions using thermogravimetric analyzer (TGA), differential scanning calorimeter (DSC), and gas chromatograph - mass spectrometer (GC-MS). The progress of the reaction was observed in situ through monitoring the release of water (mass loss and infra-red spectra) as well as the characterization of the solid residue (1H and 13C NMR, powder and single crystal XRD). The scalability of the reaction was tested with larger samples using a ventilation oven, and a microwave reactor. We found that the both dipeptides easily undergo the cyclization reaction upon mild heating, to give a stereospecific product with ~100% yield. However, the study was complicated with polymorphism displayed by the cyclic product. The solid state reaction yielded an orthorhombic form of the cyclic dipeptide, while its recrystallization produced a triclinic polymorph. The crystal structure and relative stability of the forms were investigated with XRD and DSC techniques. When the solid-state reaction was conducted at higher temperature, partial racemization and distereomerization were observed which led to new crystal structures. The degree of racemization was evaluated by a polarimeter and the NMR analysis.

Structure and stability of solid oligoglycines

Although peptides have been extensively studied within many disciplines, their solid state chemistry is not sufficiently explored. Crystalline peptide materials could present new opportunities in solid state organic synthesis as well as pharmaceutical, cosmetic and food industries. In order to facilitate these future applications, a better understanding of the relationship between the solid state structure and chemical reactivity in peptide crystals is required. In this work, a series of linear glycine oligomers was studied to see how their crystal structure and thermal reactivity changes with the chains growth. Single crystals of tetra- and pentaglycines were grown from aqueous solutions and investigated with XRD analysis (see Figure for pentaglycine). The newly collected data together with those reported previously reveal general trends likely present throughout all glycine oligomers. An antiparallel, hydrogen bound, β-sheet-like structure exists throughout the whole series and is the most stable polymorphic form in higher oligomers. Further, the thermal reactivity of this series was studied using gas chromatography – mass spectrometry (GC-MS), thermal gravimetric analysis (TGA) coupled with IR spectroscopy, differential scanning calorimetry (DSC) and bulk oven heating regimes. Finally 1H and 13C NMR and XRD analysis were used to identify key components of the thermal decomposition pathways as well as new products discovered during the thermal treatment. The results from all these studies suggest that the thermal stability of the oligomers increases with the chain length, but the decomposition pathways for the series are similar. In all cases, 2,5-diketopiperazine was formed through either condensation reactions (glycine and diglycine[1]) or depolymerisation of the peptide chain[2] in parallel with competing decomposition mechanisms. 2,5-Diketopiperazine and its derivatives are important biologically active molecules, and if this trend holds for other peptide oligomers, this solid state reaction could form a new, widely applicable synthetic method.

Co-crystals of leucyl-alanine

Although the ability of different molecules to crystallize in a single solid has been known for a long time [1], some of these materials became a subject of intense studies in the last decade due to pharmaceutical and other applications [2]. Short peptides may become a very useful class of co-crystallization agents due to their wide diversity, eco-friendliness and biocompatibility [3]. In this work, we screened the dipeptide leucyl-alanine (LA, see Figure) for the ability to form co-crystals with a variety of solid bioactive compounds. Solvent-assisted grinding of two compounds was followed by a powder X-ray diffraction test. The tested bioactive compounds were found to co-crystallize successfully with the peptide when they possessed both a hydrophobic part and strong hydrogen-bonding functionality. They were primarily derivatives of benzene, phenol, pyridine, pyrazine, quinoline and isoquinoline. Nearly all compounds with an amine or amide group formed a co-crystal, whereas most carboxylic acids did not form a new phase. For the successful combinations, single crystals were obtained when possible and studied using the single-crystal X-ray diffraction analysis. To our surprise, many of the co-crystals formed contained more than the two intended components due to the incorporation of the organic solvent and/or water. For example, one of the co-crystals studied displayed a complex hydrogen bonding framework built by four types of molecules: LA, 8-quinolinecarboxylic acid, ethanol and water in a 1:0.5:0.5:0.5 ratio.

Co-crystals and inclusion compounds of leucyl-containing dipeptides

Short peptides are ecologically friendly and non-toxic molecules, so they can be safely utilized in green chemistry processes or incorporated in pharmaceuticals and food additives. It has been shown that some dipeptides can form crystals that incorporate other molecules through intermolecular hydrogen bonding and van der Waals interactions[1]. The utilization of such dipeptides for solid state organic synthesis or storage and stabilization of bioactive molecules would be of great practical interest, but the principles that define the successful combinations are not clear. In order to identify what factors lead to complementary pairs of a dipeptide and a second component, a series of leucine-containing dipeptides was screened against 40 organic solvents and solids. Direct or solvent-assisted grinding was used followed by PXRD analysis. It was found that each dipeptide was able to form new phases with some of the utilized reactive and bioactive molecules. The Figure illustrates three experimental powder patterns in the 5-35 2θ degree range. The dipeptide leucyl-valine (1) and the second component 5-acetylsalicylamide (2) combine to form a new crystalline phase (3). After screening was complete, a series of crystallizations was performed and several crystals comprised of both a dipeptide and another molecule have been isolated and studied. A number of structural motifs were observed, although a layered architecture with the second component included in the interlayer space prevailed.