文摘
The 1H and 15N NMR spectra of several 15N-labeled pyridoxal-5'-phosphate model systems havebeen measured at low temperature in various aprotic and protic solvents of different polarity, i.e.,dichloromethane-d2, acetonitrile-d3, tetrahydrofuran-d8, freon mixture CDF3/CDClF2, and methanol. Inparticular, the 15N-labeled 5'-triisopropyl-silyl ether of N-(pyridoxylidene)-tolylamine (1a), N-(pyridoxylidene)-methylamine (2a), and the Schiff base with 15N-2-methylaspartic acid (3a) and their complexes with protondonors such as triphenylmethanol, phenol, and carboxylic acids of increasing strength were studied. Withthe use of hydrogen bond correlation techniques, the 1H/15N chemical shift and scalar coupling data couldbe associated with the geometries of the intermolecular O1H1N1 (pyridine nitrogen) and the intramolecularO2H2N2 (Schiff base) hydrogen bonds. Whereas O1H1N1 is characterized by a series of asymmetriclow-barrier hydrogen bonds, the proton in O2H2N2 faces a barrier for proton transfer of medium height.When the substituent on the Schiff base nitrogen is an aromatic ring, the shift of the proton in O1H1N1from oxygen to nitrogen has little effect on the position of the proton in the O2H2N2 hydrogen bond. Bycontrast, when the substituent on the Schiff base nitrogen is a methyl group, a proton shift from O to N inO1H1N1 drives the tautomeric equilibrium in O2H2N2 from the neutral O2-H2···N2 to the zwitterionicO2-···H2-N2+ form. This coupling is lost in aqueous solution where the intramolecular O2H2N2 hydrogenbond is broken by solute-solvent interactions. However, in methanol, which mimics hydrogen bonds tothe Schiff base in the enzyme active site, the coupling is preserved. Therefore, the reactivity of Schiff baseintermediates in pyridoxal-5'-phosphate enzymes can likely be tuned to the requirements of the reactionbeing catalyzed by differential protonation of the pyridine nitrogen.