Effect of conformation on the rate of deamidation of vancomycin in aqueoussolutions

The instability of vancomycin, a glycopeptide antibiotic, limits its shelf- life because the deamidation of its asparagine residue results in the forma tion of a zwitterion with limited aqueous solubility. Analysis of the pi-I- rate profile for vancomycin indicates that the deamidation reaction is nota bly sensitive to the ionic state of the molecule. This observation results in a hypothesis in which the ionic state of vancomycin may influence the co nformation of the molecule and therefore affect its reactivity. Two-dimensi onal nuclear magnetic resonance (NMR), homonuclear Hartmann-Hahn (HOHAHA) a nd rotating frame Overhauser enhancement spectroscopy (ROESY) information c ombined with molecular dynamic simulations were used to estimate the appare nt conformation of vancomycin in aqueous solution at pH 4 and pH 9 where th e molecule exists primarily as a monocation and monoanion, respectively. Th e apparent conformation for vancomycin at pH 4 is compact, and the proximit y of the backbone amide nitrogen to the side chain carbonyl carbon of aspar agine is favorable for the rapid formation of the cyclic imide intermediate , thus increasing its reactivity. The apparent conformation for vancomycin at pH 9, however, is expanded in comparison with the conformation at pH 4, and the increase in distance between the reacting atoms leads to slower cyc lic imide formation and thus decreased intrinsic reactivity. That cyclic im ide formation was rate limiting at both pH values was confirmed by cyclic i mide isolation and stability estimation. It becomes apparent from the analy sis of the pH-rate and conformational profiles of vancomycin that the deami dation rate of vancomycin is largely influenced by the ionization state of the N-methyl leucine nitrogen. (C) 2000 Wiley-Liss, Inc.