Glassy carbon electrodes modified by reductive electropolymerization of a thin film of poly[Ru(vbpy)(3)(2+)] or poly[Ru(vbpy)(3)(2+)/vba] (vbpy = 4-vinyl-4'-methyl-2,2'-bipyridine and vba = p-vinylbenzoic acid) were prepared. The Ru(III/II) couples for the polymer films were reversible in nonaqueous solution but were irreversible in aqueous media. The films modified with poly[Ru(vbpy)(3)(2+)] catalyzed the oxidation of aqueous guanosine 5'-monophosphate (GMP) and poly[G], producing a current enhancement in the Ru(III/II) couple for the polymer film. The catalysis was due both to electrostatic condensation of GMP and poly[G] to the Ru-modified surface and to more facile electron transfer to the Ru(III) centers in the polymer compared to the bare electrode. The presence of GMP in solution decreased the extent of decomposition of Ru(III). When single-stranded DNA containing multiple guanines was attached to the electrode modified with the poly[Ru(vbpy)(3)(2+)/vba] copolymer, enhancement of 8-13 &mgr;A for the Ru(III/II) couple was observed with 8 pmol of attached DNA. This degree of enhancement corresponds to a current efficiency of 65% based on a one-electron oxidation of guanine.

Free energies of ionization (ΔG i°) in the gas phase have been determined for the seven fluorine-substituted ruthenocenes with formulas (η5-C5Me5)(η5-C5H5 - n F n )Ru (n = 1−5) and (η5-C5H5)(η5-C5F5)Ru. In (η5-C5Me5)(η5-C5H5 - n F n )Ru (n = 1−5), each additional fluorine was found to increase the ionization free energy by 3.7 ± 0.5 kcal mol-1, and the ΔG i° difference between the two possible isomers for n = 2 and n = 3 was found to be ≤ 1 kcal mol-1. Therefore, the intrinsic effect of each fluorine substituent on ionization energy is approximately additive.

The presence of the Zn2+ ion dramatically enhances the inhibition of trypsin and tryptase by amidine-modified benzimidazole inhibitors via coordination to both the catalytically active Ser195 hydroxyl and His57 imidazole residues of the enzyme and the nitrogens of the amidine-modified benzimidazole inhibitor (Janc, J. W.; Clark, J. M.; Warne, R. L.; Elrod, K. C.; Katz, B. A.; Moore, W. R. Biochemistry 2000, 39, 4792-4800). Some new 5-amidino-2-substituted benzimidazoles were synthesized and compared to known related molecules to explore systematically the metal-mediated inhibition of bovine trypsin as a function of coordinating groups and metal ions. These compounds take advantage of the favorable interaction between the amidine group on one side of the inhibitor and the Asp189 carboxylate in the binding pocket of the enzyme. The 5-amidino-2-substituted benzimidazoles all demonstrated similar inhibition constants (Ki) of 20-50 microM in the absence of metal ions. In the presence of Zn2+, inhibition increased to varying extents, depending upon the group substituted at the 2 position of the benzimidazole. The largest increase in inhibition in the presence of Zn2+ was seen with (5-amidino-2-benzimidazolyl)-2-benzimidazolylmethane with an apparent inhibition constant (Ki') of 0.37 +/- 0.06 nM, giving a 59,000-fold increase in inhibition when Zn2+ is present. Other metal ions, including Mn2+, Sc3+, and Hg2+, also increased the inhibition by several of the benzimidazole derivatives synthesized. The compound bis(2-benzimidazolyl)methane (BBIM) was also examined because it lacks the amidine group that provides a favorable hydrogen-bonding interaction with Asp189 in the binding pocket of trypsin. In the absence of metal ions, BBIM did not have a detectable affinity for trypsin; however, in the presence of Zn2+, a Ki' of 127 +/- 3 nM was observed. This result demonstrates that an affinity for the enzyme in the absence of metal ions is not required for potent metal-mediated inhibition, greatly expanding the possibilities for metal mediation of nonmetalloenzymes.