Do proteins at low temperature behave as glasses? A single-molecule study

We have recorded long spectral diffusion trajectories from individual LH2 pigment-protein complexes from the purple bacterium Rhodobacter sphaeroides at 1.4 K. From these data, the spectral cumulants of the absorption lines of individual, protein-embedded BChl a pigments have been evaluated. It appears that the first and second cumulants cannot be described by the predictions of the well tested standard two-level system (TLS) model for spectral diffusion in glasses. The results of the present study clearly show that there is a fundamental difference between the relaxation behavior of our test protein and that of glasses.     

Understanding functional diversity and substrate specificity in haem peroxidases: what can we learn from ascorbate peroxidase?

This review summarizes the most recent advances in our understanding of the haem enzyme ascorbate peroxidase. The aim is to show how the combined applications of protein engineering, mechanistic and structural studies can be used to provide an overall picture of enzyme catalysis, and how this information can be used to provide new insight into other, more well-characterized peroxidases (in particular cytochrome c peroxidase). It contains 212 references and covers literature up to March 2003.     

Designer haem proteins: What can we learn from protein engineering?

Iron protoporphyrin(IX) is one of the most versatile and widespread pieces of catalytic machinery known in biology and is a key component of a multitude of proteins and enzymes. One of most challenging questions in this area has been to identify and understand the relationships that exist between different classes of haem proteins and to use protein engineering methods to rationalize the mechanisms by which the protein structure controls the specific chemical reactivity of the haem group. The application of this approach to the haem enzyme ascorbate peroxidase and the haem protein leghaemoglobin is discussed. © 2002 Wiley Periodicals, Inc. Heteroatom Chem 13:501–505, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.10094     

H4(+): what do we know about it?

The potential energy surface of H(4)(+) has been analyzed and stationary points and minima of intersections characterized by benchmark multireference configuration interaction calculations with basis sets as large as augmented septuble zeta. No evidence for minima other than those of the well established stable C(2v) configuration has been found. Some of the results obtained previously at a lower level of ab initio theory had to be revised.     

Probing chemical shifts of invisible states of proteins with relaxation dispersion NMR spectroscopy: how well can we do?

Carr-Purcell-Meiboom-Gill relaxation dispersion NMR spectroscopy has evolved into a powerful approach for the study of low populated, invisible conformations of biological molecules. One of the powerful features of the experiment is that chemical shift differences between the exchanging conformers can be obtained, providing structural information about invisible excited states. Through the development of new labeling approaches and NMR experiments it is now possible to measure backbone 13C(alpha) and 13CO relaxation dispersion profiles in proteins without complications from 13C-13C couplings. Such measurements are presented here, along with those that probe exchange using 15N and 1HN nuclei. A key experimental design has been the choice of an exchanging system where excited-state chemical shifts were known from independent measurement. Thus it is possible to evaluate quantitatively the accuracy of chemical shift differences obtained in dispersion experiments and to establish that in general very accurate values can be obtained. The experimental work is supplemented by computations that suggest that similarly accurate shifts can be measured in many cases for systems with exchange rates and populations that fall within the range of those that can be quantified by relaxation dispersion. The accuracy of the extracted chemical shifts opens up the possibility of obtaining quantitative structural information of invisible states of the sort that is now available from chemical shifts recorded on ground states of proteins.     

Photochemical properties of carotenoids: what can we get from the VB model?

Photo-isomerization and anti-oxidation of carotenoids have been studied for many years be-cause of their diverse roles in photobiology, photochemistry and photomedicine[1—6]. The experi-mental works revealed that the changes in the geometry between S0 (the ground state) and T1 (the first triplet state) states are very important for the two processes. Meanwhile, theoretical studies have also been carried out to investigate these processes. The polyenes have usually been used as the models for…     

Cell electrophoresis on a chip: what can we know from the changes in electrophoretic mobility?

An overview of both experimental and theoretical studies of cell electrophoresis mobility (EPM) over the past fifty years and the relevance of cell EPM measurement are presented and discussed from the viewpoint of exploring the potential use of cell EPM as an index of the biological condition of cells. Physical measurements of the optical and/or electrical properties of cells have been attracting considerable attention as noninvasive cell-evaluation methods that are essential for the future of cell-based application technologies such as cell-based drug screening and cell therapy. Cell EPM, which can be measured in a noninvasive manner by cell electrophoresis, reflects the electrical and mechanical properties of the cell surface. Although the importance of cell EPM has been underestimated for a long time, mostly owing to the technical difficulties associated with its measurement, recent improvements in measurement technology using microcapillary chips have been changing the situation: cell EPM measurement has become more reliable and faster. Recent studies using the automated microcapillary cell electrophoresis system have revealed the close correlation between cell EPM and important biological phenomena including cell cycle, apoptosis, enzymatic treatment, and immune reaction. In particular, the converged EPM distribution observed for synchronized cells has altered the conventional belief that cell EPMs vary considerably. Finding a new significance of cell EPM is likely to lead to noninvasive cell evaluation methods essential for the next-generation of cell engineering.     

Why do Congo Red,Evans Blue,and Trypan Blue differ in their complexation properties?

Congo Red, Evans Blue, and Trypan Blue dyes were evaluated in terms of their ability to form supramolecular systems in water solution. A geometric transformation set was defined to construct a supramolecular model system composed of eight dye molecules. The stability of the constructed multimolecular systems was estimated by molecular dynamics using AMBER 4.1 and DISCOVER force fields. The results essentially confirm the tendency toward self‐assembly in each case. However, Congo Red and Evans Blue were found to form stable, continuous, ribbon‐like supramolecular organizations, whereas Trypan Blue self‐assembly appeared defective; some additional deviations from planarity seem to be the main reason for the disturbance in self‐assembling. The extra rotation around the azo bonds in the Trypan Blue molecule is slightly extorted by the close proximity of sulfonic groups. This may also be the direct cause of the observed deviation from symmetry in the molecule of this dye. The results confirm that the self‐assembling capability of the compounds studied correlates with the capacity to complex proteins, supporting the idea that supramolecularity may create specific ligation properties. © 2000 John Wiley & Sons, Inc. J Comput Chem 21: 656–667, 2000     

Esterification of alcohols with acetic anhydride in Br?nsted acidic ionic liquids at room temperature

Br?nsted acidic ionic liquids were employed as a series of efficient catalysts and solvents in the esterification of alcohols with acetic anhydride at room temperature. Good yields, mild reaction conditions, short reaction time and no side reactions were achieved. The ionic liquids could be recycled easily without obvious decline in catalytic activities.     

Current trends in lead discovery: are we looking for the appropriate properties?

The new drug discovery paradigm is based on high-throughput technologies, both with respect to synthesis and screening. The progression HTS hits lead series candidate drug marketed drug appears to indicate that the probability of reaching launched status is one in a million. This has shifted the focus from good quality candidate drugs to good quality leads. We examined the current trends in lead discovery by comparing MW (molecular weight), LogP (octanol/water partition coefficient, estimated by Kowwin [17]) and LogSw (intrinsic water solubility, estimated by Wskowwin [18]) for the following categories: 62 leads and 75 drugs [11]; compounds in the development phase (I, II, III and launched), as indexed in MDDR; and compounds indexed in medicinal chemistry journals [ref. 20], categorized according to their biological activity. Comparing the distribution of the above properties, the 62 lead structures show the lowest median with respect to MW (smaller) and LogP (less hydrophobic), and the highest median with respect to LogSw (more soluble). By contrast, over 50% of the medicinal chemistry compounds with activities above 1 nanomolar have MW > 425, LogP > 4.25 and LogSw < -4.75, indicating that the reported active compounds are larger, more hydrophobic and less soluble when compared to time-tested quality leads. In the MDDR set, a progressive constraint to reduce MW and LogP, and to increase LogSw, can be observed when examining trends in the developmental sequence: phase I, II, III and launched drugs. These trends indicate that other properties besides binding affinity, e.g., solubility and hydrophobicity, need to be considered when choosing the appropriate leads.     

Direct and indirect DIET and DIMET from semiconductor and metal surfaces: what can we learn from 'toy models'?

Desorption induced by electronic transitions (DIET) and its variant DIMET (M = 'Multiple'), are among the simplest possible "reactions" of ad-species involving ultra-short lived electronically excited states at surfaces. The non-adiabatic bond-cleavage can be enforced, for example, with laser irradiation or with electrons or holes emitted from the tip of a scanning tunnelling microscope (STM). The transient creation of excited intermediates can proceed directly (localised to the adsorbate-substrate complex), or indirectly (i.e., through the substrate). To understand the basic processes, simple one-mode two-state "toy models" such as the Menzel-Gomer-Redhead (MGR) or the Antoniewicz scenarios have proven very useful in the past. We adopt and extend MGR- and Antoniewicz-type models together with numerically exact open-system density matrix theory to address a few actual problems/experiments in DI(M)ET: (1) Direct, laser-induced desorption of H(D) from Si(100) surfaces which has been realised in the continuous-wave DIET regime only recently [T. Vondrak and X.-Y. Zhu, Phys. Rev. Lett., 1999, 82, 1967], is studied and compared to so-far hypothetical femtosecond laser desorption. The possibility of controlling the reaction by shaping the laser pulses is addressed. (2) For the same system, temperature effects are studied for electron- or hole-stimulated desorption with an STM [T. C. Shen, C. Wang, G. C. Abeln, T. R. Tucker, J. W. Lyding, Ph. Avouris and R. E. Walkup, Science, 1995, 268, 1590; C. Thirstrup, M. Sakurai, T. Nakayama and K. Stokbro, Surf. Sci., 1999, 424, L329]. A modified version of Gadzuk's "sudden transition and averaging" approach is adopted which accounts for temperature dependent excited state lifetimes. (3) For photodesorption of NO from Pt(111), based on quantum dynamical simulations possible experimental tests involving static electric fields are suggested to address the relevance of the recently challenged [F. M. Zimmermann, Surf. Sci., 1997. 390, 174], "negative ion resonance" model of the Antoniewicz type.     

Interfaces between coexisting phases in polymer mixtures: What can we learn from Monte Carlo simulations?

Symmetric binary polymer mixtures are studied by Monte Carlo simulation of the bond fluctuation model, considering both interfaces between coexisting bulk phases and interfaces confined in thin films. It is found that the critical behavior of interfacial tension and width is compatible with that of the Ising model, as expected from the universality principle. In the strong segregation limit, only qualitative but not quantitative agreement with the self-consistent field (SCF) theory is found. It is argued that the SCF theory requires but for the short chains studied (N = 32 effective monomer units per chain), the limit is only reached for close to unity. Also, the effective χ-parameter decreases in the interface. It is shown that the interfacial width w does not increase by the adsorption of block copolymers as long as their areal density is still dilute (“mushroom” regime). But a broadening of interfaces does occur for thin films confined between walls at distance D, due to fluctuations that lead to for short-range forces, in agreement with experiment.     

Intramolecular charge transfer in π-conjugated oligomers: a theoretical study on the effect of temperature and oxidation state

Intramolecular charge transfer (ICT) of gaseous π-conjugated oligo-phenyleneethynylenes (OPE) induced by a homogeneous applied electric field has been theoretically investigated using a combined approach integrating molecular dynamics (MD) simulations and Perturbed Matrix Method calculations. In line with recent investigations, our results indicate the peculiar role of conformational transitions on OPE electronic properties which reflects on a strong temperature effect on ICT. Electron transfer reactions inducing chemical alteration on OPE, also taken into account in this study, revealed extremely important for explaining non-linear ICT effects and probably plays a central role in the mechanisms underlying molecular transport junctions. Our study further points out the necessity of using MD-based approach for modelling molecular electronics, even when relatively rigid molecular systems are concerned.     

2-methylterrylene in hexadecane: do we see single rotational quantum jumps of methyl groups?

We performed comparative low temperature (2-30 K) hole-burning and single molecule experiments with 2-methylterrylene with the goal to detect single rotational tunneling jumps of methyl groups. The hole-burned spectrum with its sharply structured side features which are perfectly symmetrically arranged with respect to the central hole supports the assignment to rotational tunneling transitions. However, instead of one, three clearly distinguishable methyl groups show up in the spectrum. Based on molecular mechanics simulations we attribute them to different, nearly degenerate orientations of guest molecules in one specific site of the hexadecane lattice. The frequency distribution of spontaneous jumps of single molecules reflects the features of the hole-burned spectra, although the distribution in the single molecule experiments is significantly broader. The photoinduced frequency transformation of single molecules ("single molecule photobleaching experiments") fits to the features of the hole-burned spectra, except that, surprisingly, no significant number of spectral jumps could be generated in the frequency range where the prominent narrow antiholes are observed in the hole-burned spectra.     

Looking at long molecules in solution: what happens when they are subjected to Couette flow?

Knowing the structure of a molecule is one of the keys to deducing its function in a biological system. However, many biomacromolecules are not amenable to structural characterisation by the powerful techniques often used namely NMR and X-ray diffraction because they are too large, or too flexible or simply refuse to crystallize. Long molecules such as DNA and fibrous proteins are two such classes of molecule. In this article the extent to which flow linear dichroism (LD) can be used to characterise the structure and function of such molecules is reviewed. Consideration is given to the issues of fluid dynamics and light scattering by such large molecules. A range of applications of LD are reviewed including (i) fibrous proteins with particular attention being given to actin; (ii) a far from comprehensive discussion of the use of LD for DNA and DNA-ligand systems; (iii) LD for the kinetics of restriction digestion of circular supercoiled DNA; and (iv) carbon nanotubes to illustrate that LD can be used on any long molecules with accessible absorption transitions.     

How do rotameric conformations influence the time-resolved fluorescence of tryptophan in proteins? A perspective based on molecular modeling and quantum chemistry

We discuss the dynamics of tryptophan rotamers in the context of the non-exponential fluorescence decay in proteins. The central question is: how does the ground-state conformational heterogeneity influence the time evolution of tryptophan fluorescence? This problem is examined here from the theoretical perspective. Three methods at different levels of theory, and with different scopes and computational requirements are reviewed. The Dead-end elimination method is limited to side-chain dynamics and provides an efficient way to detect the stable tryptophan rotamers in a protein. Its application to the study of heterogeneous emission characteristics is illustrated. Molecular dynamics is aimed at the full phase space of the macromolecule in solution, but must rely on classical force fields and laws of evolution. We examine to what extent the molecular mechanics paradigm yields sufficiently accurate thermodynamic results, and what are the possible kinetic implications. Finally Quantum Chemistry is the only theoretical method that allows a direct assessment of the excited states. It is necessarily restricted to small molecular systems, and thus must be used in a hybrid combination with classical methods and electrostatic models. So far understanding of the emitting state has greatly progressed as a result of these calculations, but the actual treatment of the photophysical decay processes at the quantum level has not yet really started.