NMR localization of protons in critical enzyme hydrogen bonds

Department of Chemistry, University of California, Davis, Davis, California, United States
Journal of the American Chemical Society (Impact Factor: 11.44). 09/2007; 129(31):9558-9. DOI: 10.1021/ja0728223
Source: PubMed

ABSTRACT Using N-15 NMR spectroscopy and hydrogen bond correlations, we have localized a mechanistically critical proton in aspartate aminotransferase in microcrystals and aqueous solution. It is in a H-bond between a carboxylate O of Asp222 and the pyridine nitrogen of pyridoxal-5'-phosphate. At neutral pH in water, aspartate and pyridine are unprotonated, but they share a proton in the enzyme. It is shown that such a binuclear base is typical for acid-base interactions in aprotic polar solvents. Active site H-bonds to Asp222 assist protonation of the pyridine nitrogen in the enzyme, which is considered a prerequisite for catalytic activity. We also show that acid-base behavior in enzymes should be modeled using aprotic polar solvents rather than aqueous solutions.


Available from: Peter M Tolstoy, Jun 13, 2015
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