NMR Localization of Protons in Critical Enzyme Hydrogen Bonds

详细信息    查看全文

• 作者：Shasad Sharif ; Emily Fogle ; Michael D. Toney ; Gleb S. Denisov ; Ilya G. Shenderovich ; Gerd Buntkowsky ; Peter M. Tolstoy ; Monique Chan Huot ; Hans-Heinrich Limbach
• 刊名：Journal of the American Chemical Society
• 出版年：2007
• 出版时间：August 8, 2007
• 年：2007
• 卷：129
• 期：31
• 页码：9558 - 9559
• 全文大小：411K
• 年卷期：v.129,no.31(August 8, 2007)
• ISSN：1520-5126

文摘

Using ¹⁵N 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.