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Mammalian (Clade 3) catalases utilize NADPH as a protective cofactor to prevent one-electron reduction of the central reactive intermediate Compound I (Cpd I) to the catalytically inactive Compound II (Cpd II) species by re-reduction of Cpd I to the enzyme's resting state (ferricatalase). It has long been known that ascorbate/ascorbic acid is capable of reducing Cpd I of NADPH-binding catalases to Cpd II, but the mode of this one-electron reduction had hitherto not been explored. We here demonstrate that ascorbate-mediated reduction of Cpd I, generated by addition of peroxoacetic acid to NADPH-free bovine liver catalase (BLC), requires specific binding of the ascorbate anion to the NADPH binding pocket. Ascorbate-mediated Cpd II formation was found to be suppressed by added NADPH in a concentration-dependent manner, for the achievement of complete suppression at a stoichiometric 1:1 NADPH:heme concentration ratio. Cpd I → Cpd II reduction by ascorbate was similarly inhibited by addition of NADH, NADP(+), thio-NADP(+), or NAD(+), though with 0.5-, 0.1-, 0.1-, and 0.01-fold reduced efficiencies, respectively, in agreement with the relative binding affinities of these dinucleotides. Unexpected was the observation that although Cpd II formation is not observed in the presence of NADP(+), the decay of Cpd I is slightly accelerated by ascorbate rather than retarded, leading to direct regeneration of ferricatalase. The experimental findings are supported by molecular mechanics docking computations, which show a similar binding of NADPH, NADP(+), and NADH, but not NAD(+), as found in the X-ray structure of NADPH-loaded human erythrocyte catalase. The computations suggest that two ascorbate molecules may occupy the empty NADPH pocket, preferably binding to the adenine binding site. The biological relevance of these findings is discussed.

A pyrene-functionalized cationic oligopeptide 1 efficiently binds to double-stranded DNA, as shown by different spectrophotochemical studies. Upon binding, the conformation of 1 changes from a folded to an extended form, which leads to a distinct change in the fluorescence properties. Thus, 1 functions as a molecular peptide beacon, and as it is easily taken up by cells, 1 can also be used for imaging of nucleic acids within cells., (© 2012 American Chemical Society)

X-ray structures of the 13 different monofunctional heme catalases published to date were scrutinized in order to gain insight in the mechanism by which NADPH in Clade 3 catalases may protect the reactive ferryloxo intermediate Compound I (Cpd I; por (*+)Fe (IV)O) against deactivation to the catalytically inactive intermediate Compound II (Cpd II; porFe (IV)O). Striking similarities in the molecular network of the protein subunits encompassing the heme center and the surface-bound NADPH were found for all of the Clade 3 catalases. Unique features in this region are the presence of a water molecule (W1) adjacent to the 4-vinyl group of heme and a serine residue or a second water molecule hydrogen-bonded to both W1 and the carbonyl group of a threonine-proline linkage, with the proline in van der Waals contact with the dihydronicotinamide group of NADPH. A mechanism is proposed in which a hydroxyl anion released from W1 undergoes reversible nucleophilic addition to the terminal carbon of the 4-vinyl group of Cpd I, thereby producing a neutral porphyrin pi-radical ferryloxo (HO-por (*)Fe (IV)O) species of reduced reactivity. This structure is suggested to be the elusive Cpd II' intermediate proposed in previous studies. An accompanying proton-shifting process along the hydrogen-bonded network is believed to facilitate the NADPH-mediated reduction of Cpd I to ferricatalase and to serve as a funnel for electron transfer from NADPH to the heme center to restore the catalase Fe (III) resting state. The proposed reaction paths were fully supported as chemically reasonable and energetically feasible by means of density functional theory calculations at the (U)B3LYP/6-31G* level. A particularly attractive feature of the present mechanism is that the previously discussed formation of protein-derived radicals is avoided.

Acyclic receptors containing neutral hydrogen bonding sites, such as amino-pyridine groups and a crown unit, perform effective recognition of neutral sugar molecules through multiple interactions. Receptor 1 has been shown to be a particularly effective and highly selective receptor for beta-glucopyranoside.

The cycloaddition of thiobenzophenone S-methylide to thiobenzophenone, an experimentally well-known reaction, was studied, using (U)HF/3-21G* for finding stationary points and (U)B3LYP/6-31G*//(U)HF/3-21G* single-point calculations for energies. Some optimizations were performed by (U)B3LYP/ 6-31G* to check the reliability of the calculations. The comparison of the concerted pathways and stepwise reactions via C,C-biradicals and C,S-zwitterions showed that the formation of a tetraphenyl-substituted C,C-biradical and its ring closure to 4,4,5,5-tetraphenyl-1,3-dithiolane constitutes the energetically most probable pathway of product formation, despite the fact that the regioisomeric 2,2,4,4-tetraphenyl-substituted product is more favorable by 17 kcal mol(-1). Model calculations on bond dissociation energies showed that (U)B3LYP with various basis sets overestimates radical stabilization, whereas CBS-QB3 closely reproduced experimental values. Results with the BLYP functional are similar to those with B3LYP. The consequences of the overestimation of radical stability for the cycloaddition mechanism involving biradicals are discussed. Thiobenzophenone S-methylide, if not captured by a dipolarophile, dimerizes to 2,2,3,3-tetraphenyl-1,4-dithiane. Calculation disclosed likewise a tetraphenyl-substituted C,C-biradical as intermediate.

[structure: see text] New effective, acyclic, pyridine-based receptors 1-3 show remarkable alpha/beta binding selectivity in the recognition of monosaccharides. They are able to participate in cooperative and bidentate hydrogen bonds with sugar hydroxyls as well as in CH-pi interactions with CH's of sugar molecules.