Electronic and Nuclear Magnetic Resonance Spectroscopic Features of the 1‘,4‘-Iminopyrimidine Tautomeric Form of Thiamin Diphosphate, a Novel Intermediate on Enzymes Requiring This Coenzyme^†

Appropriate compounds were synthesized to create models for the 1‘,4‘-imino tautomer of the 4‘-aminopyrimidine ring of thiamin diphosphate recently found to exist on the pathway of enzymatic reactions requiring this cofactor [Jordan, F., and Nemeria, N. S. (2005) Bioorg. Chem. 33, 190−215]. The N1-methyl-4-aminopyrimidinium compounds synthesized on treatment with a strong base produce the 1,4-imino tautomer whose UV spectrum indicates a maximum between 300 and 320 nm, depending on the absence or presence of a methyl group at the 4-amino nitrogen. The λ_max found is in the same wavelength range as the positive circular dichroism band observed on several enzymes and showed a very strong dependence on solvent dielectric constant. To help with the ¹⁵N chemical shift assignments, the model compounds were specifically labeled with ¹⁵N at the amino nitrogen atom. The chemical shift of the amino nitrogen was deshielded by N1-methylation and then dramatically further deshielded by more than 100 ppm on formation of the 1,4-iminopyrimidine tautomer. Both the UV spectroscopic values and the ¹⁵N chemical shift for the 1,4-iminopyrimidine tautomer should serve as useful guides to the assignment of enzyme-bound signals.