Selenium Atom-Specific Mutagenesis (SAM) for Crystallography, DNA Nanostructure Design, and Other Applications - DNA Nanotechnology: From Structure to Function - page 43

Scheme 3.8 Synthesis of 2-Se-T phosphoramidite and DNAs. Reagents and conditions: ( a )
DMTr-Cl, pyridine, DMAP, rt; ( b )DBU,DMF,CH 3 I; ( c )Se,NaBH 4 , EtOH; ( d ) I-CH 2 CH 2 CN,
i -Pr 2 NEt, CH 2 Cl 2 ;( e )( i -Pr 2 N) 2 P(Cl)OCH 2 CH 2 CN, ( i -Pr) 2 NEt, CH 2 Cl 2 ;( f ) Solid-phase synthesis
was investigated. Though the 2-Se-derivatized nucleoside was synthesized over
three decades ago [ 80 , 81 ], 2-Se-T was never incorporated into DNA due to the
synthetic difficulties in the Se functionality protection and deprotection. Recently,
we have developed a novel synthetic route (Scheme 3.8 ) to incorporate the selenium
functionality by displacing the 2-Se moiety with the selenium functionality [ 18 ],
which is a convenient and efficient strategy for the selenium incorporation. The key
selenization reaction is accomplished by displacing the 2-methylsulfide activating
group with sodium hydrogen selenide. This reaction is also called the Huang
reaction. Later, the 2-Se-T phosphoramidite was successfully synthesized and
incorporated into DNA for studying the base-pair recognition and duplex structure.
This novel synthesis of 2-Se-T utilizes 2-methylthio-T as an active intermediate,
followed by the methylation and the selenizing reaction (Huang reaction) with
NaSeH to give 2-seleno-T (Scheme 3.8 ). To ensure the stability of the selenium
functionality during solid-phase synthesis, cyanoethyl group is used to protect it,
preventing the deselenization in solid-phase synthesis cycle. Biophysical experi-
ments of the 2-Se-T-DNAs have demonstrated largely increased base-pair specificity
by destabilizing T/G (wobble pair) and T/C pairs. Although the thymidine 2-exo-
position is not involved in Watson-Crick base pairing (or hydrogen bonding), it
is involved in the wobble T/G pairing. The 2-Se moiety discriminates against the
wobble pairing via the steric and electronic effects of the selenium atom. The
crystallographic characterization also suggests that 2-Se-T has higher base-pairing
specificity over native T (Fig. 3.6 ). This result was also later supported by the
computational study [ 82 ].