Synthesis of Deoxypseudouridine 5'-Triphosphate Bearing the Photoremovable Protecting Group at the 1 Position Capable of Enzymatic Incorporation to DNA.

Enzymatic incorporation of deoxynucleoside 5'-triphosphate bearing the photocleavable protecting group is a useful method for the preparation of photocaged oligodeoxynucleotides. Here, we describe the synthesis of new photocaged deoxynucleoside triphosphates 1-(2-nitrobenzyl)-deoxypseudouridine triphosphate (dΨTP) and 1-(6-nitropiperonyloxymethyl)-deoxypseudouridine triphosphate (dΨTP). We successfully synthesized dΨTP and dΨTP and applied them to enzymatic synthesis of photocaged oligonucleotides.

Deoxynucleoside Triphosphate Containing Pyridazin-3-one Aglycon as a Thymidine Triphosphate Substitute for Primer Extension and Chain Elongation by Klenow Fragments.

Deoxynucleoside 5'-triphosphate was synthesized with 3-oxo-2 H-pyridazin-6-yl (Pz)-a uracil analogue lacking a 2-keto group-as the nucleobase. Theoretical analyses and hybridization experiments indicated that Pz recognizes adenine (A) for formation of a Watson-Crick base pair. Primer extension reactions using nucleoside 5'-triphosphate and the Klenow fragment revealed that the synthetic nucleoside 5'-triphosphate was incorporated into the 3' end of the primer through recognition of A in the template strand.

Synthesis of photocaged 6-O-(2-nitrobenzyl)guanosine and 4-O-(2-nitrobenzyl) uridine triphosphates for photocontrol of the RNA transcription reaction.

6-O-(2-Nitrobenzyl)guanosine and 4-O-(2-nitrobenzyl)uridine triphosphates (GTP, UTP) were synthesized, and their biochemical and photophysical properties were evaluated. We synthesized UTP using the canonical triphosphate synthesis method and GTP from 2',3'-O-TBDMS guanosine via a triphosphate synthesis method by utilizing mild acidic desilylation conditions. Deprotection of the nitrobenzyl group in GTP and UTP proceeded within 60s by UV irradiation at 365nm.

Photo-controlled binding of MutS to photo-caged DNA duplexes incorporating 4-O-(2-nitrobenzyl) or 4-O-[2-(2-nitrophenyl)propyl]thymidine.

Mismatch binding protein MutS binding to bulge structure in DNA duplexes was controlled by UV irradiation. 4-O-(2-Nitrobenzyl)thymidine or 4-O-[2-(2-nitrophenyl)propyl]thymidine was incorporated into DNA duplexes a bulged position. The MutS did not bind to the caged DNA duplexes but bound after removing the 2-nitrobenzyl or 2-(2-nitrophenyl)propyl group by photo-irradiation.