Editorial of the PCCP themed issue on “Physical Chemistry for Life Sciences”

Probably the most influential idea of the 19th century's French philosopher Auguste Comte was his classification of sciences (1854), establishing a hierarchy with mathematics at the bottom and sociology at the top, and physics, chemistry and life sciences (in ascending order) in the center. While the research target gets more and more complex going up the hierarchy, the methods become more and more detailed going down. Another, equally important, aspect of Comte's classification is that gaining knowledge in a science higher in the hierarchy depends on thoroughly mastering the sciences that occur below.

The topic of this special issue, Physical Chemistry for Life Sciences, immediately reminds us of Comte's hierarchy, since the three sciences in its center form the name of the topic. This is not accidental. A ubiquitous feature of this timely topic is the application of (e.g. spectrometric or spectroscopic) methods which were originally developed for the investigation of small molecules but are now applied to increasingly complex biomolecules like proteins, ion channels, membranes etc. or even entire cells. This development is supported by seeking answers for new questions that do not exist in small-molecule chemistry but may be of clinical relevance. On the other hand, and this is related to the second aspect of Comte's work mentioned above, a living cell cannot be completely understood without knowing in detail how rather small molecular aggregates inside the cell really operate. Despite their complex biological environment, their function can be attributed to fundamental intermolecular interactions such as hydrogen bonds or dispersion, which are important driving forces for protein folding and govern the binding of active agents to biomolecular targets.

The toolbox of physical chemistry is well-equipped to address these questions: the enormous progress in physical chemistry with regard to the development of new and the refinement of established experimental and theoretical methods supports research in life sciences in a variety of ways. Among the experimental methods, mass spectrometry, spectroscopy and microscopy play a crucial role for all disciplines of life sciences. Only some general aspects can be addressed in this article, which (including its references) should be seen in context with the forthcoming Bunsentagung 2017 and the current themed issue of PCCP published in this context.

Some methodological developments like matrix-assisted laser desorption/ionisation (MALDI) (e.g.ref. 1 and 2) and electrospray ionisation (ESI) (e.g.ref. 3 and 4), in combination with different methods to transfer energy to a molecule and thus induce fragmentation, paved the way for mass spectrometry to become a standard analytical technique in proteomics.⁵ Particularly, the developments during the last few years enabled the analysis of e.g. the entire yeast proteome within a few hours, and may even give answers to questions of clinical relevance.⁵ In this themed issue a mass spectrometric contribution from the group of Beyer [DOI: 10.1039/C6CP08436B] investigates the fragmentation of the neuropeptide leucine enkephalin induced by CID (collision-induced dissociation) or UV laser irradiation of variable wavelength (laser induced dissociation, LID). These methods are complementary since UV irradiation delivers the energy in large “packets” while in CID the internal energy slowly increases during a number of collisions. As a consequence CID prefers dissociation pathways via “low-lying tight transition states” whereas LID favours dissociation via “high-lying loose transition states”.

Despite the analytical power of mass spectrometry, information on the structure and functionality of biological molecules or whole biological processes is preferentially drawn from spectroscopic and microscopic investigations. A variety of spectroscopic techniques are available which cover different sizes of molecular systems, aggregate states and the time resolution. To gain fundamental understanding of complex biological systems, a bottom-up approach can start with the analysis of single molecular components of such a system. For this purpose different laser spectroscopic methods (frequently combined with mass spectrometry) are applied to isolated neutral and charged species in molecular beams (e.g.ref. 6–10) or ion traps (e.g.ref. 11–15). They give insight into fundamental molecular interactions like hydrogen bonds, dispersion or electrostatic stabilization, which are important driving forces for protein folding and play a key role in rational drug design for specific biomolecular targets. Several contributions in this themed issue address the first step of this bottom-up approach.

The report from the groups of Dopfer and Fujii [DOI: 10.1039/C6CP08553A] investigates the conformers of gas-phase protonated glutamic acid via IR(M)PD spectroscopy (Infrared Multi Photon Dissociation). Since even the smallest biomolecules like amino acids have a rich conformation landscape, extended conformational analyses are performed with density functional methods, and calculated vibrational spectra for the most stable conformers found in the calculations are compared with the experimental IR(M)PD spectra. This combination of experiment and theory then revealed that at low temperature two conformers are present. As an alternative to IR(M)PD, IR/UV dip spectra can provide the vibrational frequencies of gas phase molecules. This approach was used by the groups of Jouvet and Fujii to investigate the protonated neurotransmitter noradrenaline [DOI: 10.1039/C6CP08426E]. As in the contribution discussed before, a comparison with calculated vibrational frequencies was performed and revealed five conformers, also higher energy ones which originate from solution conditions and are kinetically trapped in the cooling process. The knowledge of the neurotransmitter's conformational landscape is of scientific interest because upon binding of noradrenaline to its receptor, a single conformation is mapped out.

Beyond charged systems, neutral, biologically relevant molecules were also analysed. Among them is N-methyl acetamide, whose dimer and trimer are considered to be the smallest model system for N–H/C=O hydrogen-bonding interactions determining the secondary structure of peptides. Unfortunately, the experimental characterisation of this seemingly simple system was far from being complete, and the contribution from the group of Suhm demonstrates that earlier spectroscopic assignments have to be corrected [DOI: 10.1039/C6CP07989J]. This has been achieved using Raman spectroscopy in combination with improved quantum chemical calculations.

The contribution of the Gerhards group aims to identify the most stable conformers of cyclic tetrapeptides cyclo[L-Tyr(Me)-D-Pro-L-X-D-Pro] (X = Ala, Glu(Me), Tyr(Me)) in the gas phase by recording IR/R2PI (Infrared/Resonant 2 Photon Ionisation) spectra in combination with quantum chemical calculations [DOI: 10.1039/C6CP08696A]. It was found that cyclotetrapeptides generally form doubly hydrogen-bonded structures but if this peptide contains an amino acid like glutamic acid with a flexible and H-bond accepting side chain a new energetically favourable conformation with a different hydrogen bonding pattern is additionally observed.

The spectroscopic methods addressed above are mostly based on lasers as light sources, whose application yield a variety of non-linear effects. However, the introduction of methods directly taking advantage of these non-linear effects substantially increased the pool of physico-chemical methods (e.g.ref. 16 and 17). One of the non-linear spectroscopic methods addressed in this themed issue is vibrational sum frequency generation (VSFG) which is an important method to investigate (monolayer) adsorption on interfaces: since the contributions from randomly oriented molecules in the bulk compensate each other, the recorded spectrum only arises from oriented molecules in the vicinity of the interface. In their contribution, Meister, Paananen and Bakker analyse (in dependence of altering pH-values) highly-ordered hydrophobin-containing protein films at the water–air interface via conventional and heterodyne-detected VSFG. In general the interference of the broad O–H band of bulk water with protein N–H bands can lead to misinterpretations in the conventional VSFG spectra [DOI: 10.1039/C6CP08325K]. However, the authors manage to analyse this potential source of misinterpretation of VSFG spectra using heterodyne-detected VSFG.

The structure elucidation of macromolecules like proteins, lipids, DNA and RNA as well as macromolecular complexes represents a domain of NMR and EPR spectroscopy. The latter method, attaching covalently bound paramagnetic spin labels to the macromolecule, allows information to be obtained on inter- and intramolecular distances and consequently on structures and possible conformers. The application and further refinement of pulsed high-field EPR (with internally flexible spin labels) now provides precise information on the structure of macromolecules and is able to map out their full conformational space.^18,19 A further method frequently applied to macromolecules like proteins is static and time-resolved UV/VIS absorption and fluorescence spectroscopy in solution. By applying these techniques the focus is frequently on the excited state dynamical behaviour of the molecule including processes like energy transfer. In this context the group of Dick reports on their investigations of LOV domains, the photosensory units of many proteins, which normally undergo an adduct formation in the first steps of their domain signaling. However, these investigations [DOI: 10.1039/C6CP08370F] demonstrated that in special LOV1 mutants the adduct formation is replaced by an electron transfer (photoreduction of the flavin chromophore). Having unravelled this pathway in detail paves the way for the design of new artificial photosensors, and also calls for the investigation of this single-point mutation in other LOV domains.

The phenomenon of fluorescence is nowadays applied and further developed in different spectroscopic and microscopic techniques which are even able to visualise cellular structures and whole cells. Specific methods (e.g. fluorescence correlation spectroscopy, FCS) even go beyond a pure visualisation of cellular structures and molecular distributions within cells. They are for example able to determine the dynamics and interactions of single molecules in the complex environment of a living cell (e.g.ref. 20). The application of FCS in combination with FRET (Förster Resonance Energy Transfer) based techniques e.g. allows conclusions to be drawn about the size and folding of proteins. In this year's themed issue Ebbinghaus and coworkers apply in-cell FRET microscopy to study temperature-dependent conformational dynamics and self-association of different variants of the Huntingtin exon 1 protein [DOI: 10.1039/C6CP08167C]. For the hereditary, dementia causing, Huntington's disease it is known that mutation in the Huntingtin gene affects artificially long poly-Gln chains in this Huntingtin protein leading to aggregation and intracellular inclusion bodies. As in the case of in vitro experiments, a nucleated growth mechanism is observed which means the aggregated species can act as a kind of catalytic centre to trigger further self-association. For the application of fluorescence-based methods in general, the investigated biomolecule must either intrinsically contain emitting chromophores or special fluorescence markers must be introduced. In that context fluorescent nanodiamonds can also be of interest since modified nanodiamonds with colour centers are very stable sensors and can be incorporated into cells but their uptake strongly depends on size and shape. Thus, the group of Schirhagl investigated the shape and crystal orientation of such nanodiamonds using SEM (Scanning Electron Microscopy) and AFM (Atomic Force Microscopy) [DOI: 10.1039/C6CP07431F].

Beyond the application of (laser) radiation for analytical purposes, gaining insight into biological processes on a molecular level, radiation can also be directly applied as part of a medical therapy. The holy grail of cancer therapy is a method that selectively destroys cancer cells but does little harm to the rest of the body. To this end, one needs a warhead that can be positioned close to cancer cells and which, when triggered, exerts quite local damage. In photothermal therapy, gold nanoparticles are a promising candidate for such a warhead because they bind to DNA and can absorb a near-infrared laser pulse (which has a sufficient penetration depth into body tissue). The energy absorbed is then converted to heat that produces the desired local damage. The contribution of Schürmann and Bald investigates what happens to the gold nanoparticles in a therapeutic sequence that contains thousands of laser shots, as well as the kinetics of the disintegration of DNA bases surrounding the nanoparticle [DOI: 10.1039/C6CP08433H].

A further important aspect in medical therapy is knowledge about the binding of drugs to proteins. From the thermodynamic point of view this process is, in most cases, virtually thermoneutral, such that the relative importance of enthalpic and entropic contributions to the driving force is very difficult to assess. Schäfer and coworkers use both, isothermal titration calorimetry experiments and molecular dynamics simulations to study the binding of p-aminobenzamide cation to trypsin in various water/methanol mixtures [DOI: 10.1039/C6CP07899K]. The experiments show that the binding free energy only weakly depends on solvent composition, which is the consequence of strong enthalpy/entropy compensation. The microscopic origin of this behaviour is explained by the molecular dynamics simulations.

From the large pool of available physico-chemical techniques, only a few being related to the contributions of the Bunsentagung 2017 or to the contributions to this themed issue were mentioned above. Despite the enormous capabilities of these techniques a comparison with corresponding theoretical studies is indispensable to interpret and support experimental findings and thus to gain insight into the processes of biological systems on a molecular level. Depending on the size of the investigated system, advanced ab initio and DFT-based methods (cf. e.g.ref. 6–15) or molecular dynamics simulations are established. The latter even allow the analysis of large biological entities (proteins, ion channels, membranes, etc., e.g.ref. 21 and 22).

In conclusion, taking all of these diverse contributions addressed above into account, this themed issue convincingly demonstrates how physico-chemical methods originally developed to understand chemistry now have matured to a level where they can tackle questions in the realm of life science by looking at what is going on in very fine detail. However, there is not only value in the new insight gained by applying physico-chemical methods to life science questions, but one can also see the potential for methods that can manipulate individual molecules and will sooner or later be applicable in clinical therapy.

At the end of this editorial we would like to thank all of the authors for their valuable contributions.

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