Gert van der Zwan’s research while affiliated with Vrije Universiteit Amsterdam and other places

The paper mentioned in the title contains inaccuracies and misleading interpretations that seriously misrepresent basic knowledge about tautomerism in azo dyes. Therefore, we aim in this Letter to correct the most obvious mistakes, to address the misrepresentation of the main issues of tautomerism, and to give a better understanding of the basic principles of the tautomerisation process.

Simple models, in particular those based on Onsager's point dipole in a spherical cavity embedded in a polarizable medium, played an important role in understanding equilibrium and dynamical interaction between solvent polarization and classical reaction kinetics. After a brief review of some crucial results, and an application to the interaction of fluctuating polarization field on a classical dipolar oscillator, I investigate the applicability of a similar simple model to quantum-classical interactions. Interesting differences with a fully classical system occur in equilibrium situations, and some fundamental problems with commonly used dynamical approaches are indicated.

Binding of the antihistamine drug brompheniramine (BPA) to human serum albumin (HSA) is studied by measuring quenching of the fluorescence and room temperature phosphorescence (RTP) of tryptophan. The modified Stern-Volmer equation was used to derive association constants and accessible fractions from the steady-state fluorescence data. Decay associated spectra (DAS) revealed three tryptophan fluorescence lifetimes, indicating the presence of three HSA conformations. BPA causes mainly static quenching of the long-living, solvent-exposed conformer. RTP spectra and lifetimes, recorded under deoxygenated conditions in the presence of 0.2 M KI, provided additional kinetic information about the HSA-BPA interactions. Fluorescence DAS that were also recorded in the presence of 0.2 M KI revealed that the solvent-exposed conformer is the major contributor to the RTP signal. The phosphorescence quenching is mostly dynamic at pH 7 and mostly static at pH 9, presumably related to the protonation state of the alkylamino chain of BPA. This provides direct insight into the binding mode of the antihistamine drug, as well as kinetic information at both the nanosecond and the millisecond time scales.

The Escherichia coli DNA repair enzyme AlkB is a 2-oxoglutarate (2OG)-dependent Fe(2+) binding dioxygenase that removes methyl lesions from DNA and RNA. To date, nine human AlkB homologues are known: ABH1 to ABH8 and the obesity-related FTO. Similar to AlkB, these homologues exert their activity on nucleic acids, although for some homologues the biological substrate remains to be identified. 2OG dioxygenases require binding of the cofactors Fe(2+) and 2OG in the active site to form a catalytically competent complex. We present a structural analysis of AlkB using NMR, fluorescence, and CD spectroscopy to show that AlkB is a dynamic protein exhibiting different folding states. In the absence of the cofactors Fe(2+) and 2OG, apoAlkB is a highly dynamic protein. Binding of either Fe(2+) or 2OG alone does not significantly affect the protein dynamics. Formation of a fully folded and catalytically competent holoAlkB complex only occurs when both 2OG and Fe(2+) are bound. These findings provide the first insights into protein folding of 2OG-dependent dioxygenases. A role for protein dynamics in the incorporation of the metal cofactor is discussed.

The spectroscopy and kinetics of the tyrosinase catalyzed trans-resveratrol oxidation were investigated by measuring both UV-vis absorption spectra over the 200-500 nm range and Raman spectra over the 600-1800 cm(-1) region. Room temperature UV-vis absorption spectra, as a function of time, showed the presence of two isosbestic points located at λ(1) = 270 nm and λ(2) = 345.5 nm delimiting two different regions: the reactant region around 300 nm, where the absorption decreased with time, and the product region over the low wavelength (λ < 260 nm) and high wavelength (λ > 390 nm) wavelength zone in which the absorption increased with time until, in both cases, constant values were achieved. A first-order kinetics was deduced with a rate coefficient of k(1) = (0.10 ± 0.001) min(-1), which turned out to be independent of substrate concentration over the 50-5 μM range; a feature that was rationalized by invoking the limiting case of the Michaelis-Menten scheme appropriate for substrate concentration much lower than the respective Michaelis constant. The observation of the distinct resonance enhanced Raman lines, specifically those peaking at 830 cm(-1), 753 cm(-1), and 642 cm(-1) together with their time evolution, permitted us to gain insight into some crucial features and steps of the catalytic reaction. Namely, that the formation of the so-called trans-resveratrol and tyrosinase (S)P complex with its O-O bridge plays a crucial role in the first steps of this enzymatic reaction and that the hydroxylation of the ortho C-H bond of the trans-resveratrol OH group occurs after O-O bond cleavage in the tyrosinase active site. The present study makes clear that a class of potential inhibitors of tyrosinase can be found in compounds able to bind the two Cu (II) ions of the enzyme bidentate form.

UV resonance Raman spectroscopy was used to characterize the binding of three first-generation histamine H(1) receptor antagonists-tripelennamine (TRP), mepyramine (MEP), and brompheniramine (BPA)-to human serum albumin (HSA) at pH 7.2 and pH 9.0. Binding constants differ at these pH values, which can be ascribed to the different extent of protonation of the ethylamino side chain of the ligands. We have recently shown [Tardioli et al. J. Raman Spectrosc. 2011, 42, 1016-1024] that for the solution conformation of TRP and MEP the side chain plays an important role by allowing an internal hydrogen bond with the aminopyridine nitrogen in TRP and MEP. Results presented in this paper suggest that the existence of such molecular structures has serious biological significance on the binding affinity of those ligands to HSA. At pH 7.2, only the stretched conformers of protonated TRP and MEP bind in HSA binding site I. Using UV absorption data, we derived binding constants for the neutral and protonated forms of TRP to HSA. The neutral species seems to be conjugated to a positive group of the protein, affecting both the tryptophan W214 and some of the tyrosine (Y) vibrations. BPA, for which the structure with an intramolecular hydrogen bonded side chain is not possible, is H bound to the indole ring nitrogen of W214, of which the side chain rotates over a certain angle to accommodate the drug in site I. We propose that the protonated BPA is also bound in site I, where the Y150 residue stabilizes the presence of this compound in the binding pocket. No spectroscopic evidence was found for conformational changes of the protein affecting the spectroscopic properties of W and Y in this pH range.

A paper by Amat et al. [2] reported that the ATP-driven oxidation of luciferin to electronically excited oxyluciferin catalyzed by luciferase was accelerated when ATP was priorly irradiated at non-resonant optical frequencies (NROF) at 635 and 830nm. In another paper by Amat et al. [3], increased fluorescence intensities of ATP–Mg complexes, which showed lower fluorescence than ATP when excited at 260nm, were reported in consequence of concomitant NROF irradiation (i.e., 655 and 830nm). It was postulated that NROF-induced electric field changes may alter the charge distribution in ATP's phosphate chain, resulting in lowering of the activation energy of its terminal phosphate. Here we use spectrofluorometry to further investigate this hypothesis. The effect of NROF (at 632.8nm) on the intrinsic fluorescence of non-complexed and Mg-chelated ATP in aqueous solution and the influence of NROF (514.5nm and 632.8nm) on the rate of the luciferin–luciferase reaction was studied. We found that neither the intrinsic fluorescence of ATP nor its biochemical behavior during the firefly luciferin–luciferase reaction was affected by laser irradiation with NROF. Consequently, no evidence was found supporting the postulation that NROF-induced alternations on the charge distribution of the phosphate chain affect the reactivity of ATP.