Torelli Andrew T, Spitale Robert C, Krucinska Jolanta, Wedekind Joseph E
Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, 601 Elmwood Avenue Box 712, Rochester, New York 14642, USA.
Biochem Biophys Res Commun. 2008 Jun 20;371(1):154-8. doi: 10.1016/j.bbrc.2008.04.036. Epub 2008 Apr 16.
Reaction-intermediate analogs have been used to understand how phosphoryl transfer enzymes promote catalysis. Herein we report the first structure of a small ribozyme crystallized with a 3'-OH, 2',5'-linkage in lieu of the normal phosphodiester substrate. The new structure shares features of the reaction coordinate exhibited in prior ribozyme structures including a vanadate complex that mimicked the oxyphosphorane transition state. As such, the structure exhibits reaction-intermediate traits that allow direct comparison of stabilizing interactions to the 3'-OH, 2',5'-linkage contributed by the RNA enzyme and its protein counterpart, ribonuclease. Clear similarities are observed between the respective structures including hydrogen bonds to the non-bridging oxygens of the scissile phosphate. Other commonalities include carefully poised water molecules that may alleviate charge build-up in the transition state and placement of a positive charge near the leaving group. The advantages of 2',5'-linkages to investigate phosphoryl-transfer reactions are discussed, and argue for their expanded use in structural studies.
反应中间体类似物已被用于理解磷酸转移酶如何促进催化作用。在此，我们报告了首个与3'-OH、2',5'-连接键结晶的小核酶结构，该连接键替代了正常的磷酸二酯底物。新结构具有先前核酶结构中展现出的反应坐标特征，包括模拟氧磷杂环戊烷过渡态的钒酸盐复合物。因此，该结构展现出反应中间体特征，能够直接比较RNA酶及其蛋白质对应物核糖核酸酶对3'-OH、2',5'-连接键的稳定相互作用。在各自的结构之间观察到明显的相似性，包括与可裂磷酸酯的非桥连氧形成的氢键。其他共同之处包括精心定位的水分子，其可能减轻过渡态中的电荷积累，以及在离去基团附近放置正电荷。讨论了2',5'-连接键在研究磷酸转移反应中的优势，并主张在结构研究中扩大其应用。
