Oxygen Replacement with Selenium at the Thymidine 4-Position for the Se Base Pairing and Crystal Structure Studies

Abstract

The T−A and C−G base pairing and stacking allow the formation of the stable DNA duplex structure for genetic information storage, transcription, and replication. To replace the oxygen of the nucleotide nucleobases with selenium for the studies of the base-pair recognition, the duplex stability, and the nuclei acid crystal structures, we have synthesized for the first time the 4-Se thymidine phosphoramidite and incorporated it into oligonucleotides via solid-phase synthesis with high coupling yield (99%). The Se modification on the nucleobase is relatively stable under the elevated temperature. Using the dUSe (2‘-Se-dU) to facilitate the crystallization, we have successfully crystallized the DNA containing the 4-Se−T substitution and determined its structure at 1.50 Å resolution. The UV-melting and X-ray crystal structure studies have indicated that the Se substitution on the nucleobase does not cause a significant structure perturbation, the large Se atom on the thymine can be successfully accommodated by the DNA duplex, and the Se-mediated hydrogen bond (longer than the usual hydrogen bond) is formed within the modified T−A base pair. In addition, the Se derivatization on the nucleobases further facilitates X-ray crystal structure determination of nucleic acids and their protein complexes via Se MAD phasing.

Facile synthesis of nucleoside 5′-(α-P-seleno)-triphosphates and phosphoroselenoate RNA transcription

“…Inspired by the convenient Se-derivatization of proteins, since 1998, Huang's research group has pioneered and developed the selenium derivatization of nucleic acids for X-ray crystallography (Carrasco et al 2001;Caton-Williams and Huang 2008;. We found that the selenium-derivatized RNAs and DNAs are stable and that the selenium derivatizations do not cause significant perturbation Salon et al 2007Salon et al , 2010Sheng et al 2007Sheng et al , 2011Caton-Williams and Huang 2008;Olieric et al 2009;Hassan et al 2010).…”

2-Selenouridine triphosphate synthesis and Se-RNA transcription

“…In order to minimize by-product formation, the Se-nucleobase modifications are normally protected during chemical synthesis (Salon et al 2007(Salon et al , 2008CatonWilliams and Huang 2008b;Hassan et al 2010;Sun et al 2012). Since the 2-seleno-modification on uridine is naturally occurring, we decided to directly explore its compatibility with chemical synthesis.…”

Synthesis and DNA‐Polymerase Incorporation of Colored 4‐Selenothymidine Triphosphate for Polymerase Recognition and DNA Visualization

“…[4] We have now discovered colored 4-Se-thymidine 5'-triphosphate ( Se TTP; Figure 1). We formed yellow Se TTP from colorless native TTP by altering a single atom and observed the recognition and incorporation of Se TTP by DNA polymerase.…”