Small molecules that behave like helical springs
Image: Prof. Ivan Huc
The sequential addition of amino acids during protein synthesis in biological cells initially gives rise to linear molecules. But most proteins become biologically active only after the linear chain has folded into a defined three-dimensional conformation. Local folding interactions therefore play a vital role in biological systems. For example, the structural protein elastin imparts the elasticity that allows large blood vessels to expand and contract. The highly folded and flexible titin protein, which is found in muscle, acts rather like a molecular rubber band, while proteins called collagens are responsible for both the stiffness and elasticity of fibrous tissues. Elastin, titin and collagens all adopt conformations that are largely comprised of spirally wound, alpha-helical structures. However, generally speaking, it is difficult to predict how proteins will fold, owing to the number of factors that are potentially involved in the process.
This is why Professor Ivan Huc in LMU‘s Department of Pharmacy studies aspects of molecular self-organization with the aid of model compounds called foldamers. Foldamers are synthetic polymers with a defined geometry, but they are considerably smaller than proteins, and have a distinct chemical composition. – And Huc doesn’t just design and synthesize foldamers, he explores the mechanical properties of the products.
He works with 8-hydroxyquinoline-2-carboxylic acid, rather than amino acids, as a building block, linking them together to form what are called oligoamides, which serve as models for protein folding. Oligoamides spontaneously form helices in which the quinoline rings are stacked on top of one another. So the resulting structures are clearly defined.
In his latest project, Huc turned to a technique known as single-molecule force spectroscopy to characterize the mechanical properties of oligoamides of different lengths. The method was in part developed by Professor Hermann E. Gaub, a member of the Faculty of Physics at LMU (although Gaub was not involved in the study). In such experiments, dispersed single molecules are chemically attached to a substrate. The other end of each polymer is modified in such a way that it can be captured by the minuscule tip of a flexible metal cantilever, which can be used to exert a force on the molecule. If the target adopts a helical structure in its resting state, the applied force will pull the helix apart. “Both the coupling of the tiny object to the device and the measurements of transitions with minute amplitudes are difficult to perform,” Huc explains, “and very few such experiments have successfully carried out on small molecules.” The method is more often employed to investigate the mechanics of large proteins.
Back to the original form within less than 10 microseconds
In collaboration with single molecule force spectroscopy expert Prof. Anne-Sophie Duwez, at the University of Liège in Belgium, Huc designed his molecules specifically for these experiments. “In order to study their mechanical properties, we used single-molecule force spectroscopy on oligoamides with different lengths,” he says. “We discovered that, just like mechanical springs, helical oligoamides with nanometer dimensions possess exceptional elasticity – in contrast to the behavior of most naturally occurring biopolymers.” Unlike Huc’s helical oligoamides, when large proteins are extended, they do not always quickly return to their original form upon release of the tension.
Huc and colleagues went on to show that, when exposed to stronger forces, these helices can be extended by as much as 3.8-fold relative to their starting length. As soon as the applied force is released, they return to their original form within less than 10 microseconds. Thus, as in the case of a metal spring, stretching of the molecule loads it with mechanical energy, which is rapidly restituted when the tension is released.
“The experiments show that reduced size need not impair the mechanical characteristics,” says Huc. “Our molecules, which have nanometer dimensions, are among the most robust and well-behaved springs that have ever been described.” However, they do differ in some respects from their more familiar, everyday counterparts. For example, the more a macroscopic spring is extended, the greater its resistance becomes. In the case of oligoamides, the resistance remains constant as the molecules are stretched.
And what comes next? Huc speculates that molecular springs could form the basis for the design of new classes of synthetic elastomers with customized mechanical properties.