Synthesis of 4-Thiouridine, 6-Thioinosine, and 6-Thioguanosine 3‘,5‘-O-Bisphosphates as Donor Molecules for RNA Ligation and Their Application to the Synthesis of Photoactivatable TMG-Capped U1 snRNA Fragments

Abstract

4-Thiouridine, 6-thioguanosine, and 6-thioinosine 3',5'-bisphosphates (9, 20, and 28) were synthesized in good yields by considerably improved methods. In the former two compounds, uridine and 2-N-phenylacetylguanosine were converted via transient O-trimethylsilylation to the corresponding 4- and 6-O-benzenesulfonyl intermediates (2 and 13), which, in turn, were allowed to react with 2-cyanoethanethiol in the presence of N-methylpyrrolidine to give 4-thiouridine (3) and 2-N-phenylacetyl-6-thioguanosine derivatives (14), respectively. In situ dimethoxytritylation of these thionucleoside derivatives gave the 5'-masked products 4 and 15 in high overall yields from 1 and 11. 6-S-(2-Cyanoethyl)-5'-O-(4,4'-dimethoxytrityl)-6-thioinosine (23) was synthesized via substitution of the 5'-O-tritylated 6-chloropurine riboside derivative 22 with 2-cyanoethanethiol. These S-(2-cyanoethyl)thionucleosides were converted to the 2'-O-(tert-butyldimethylsilyl)ribonucleoside 3'-phosphoramidite derivatives 7, 18, and 26 or 3',5'-bisphosphate derivatives 8, 19, and 27. Treatment of 8, 19, and 27 with DBU gave thionucleoside 3',5'-bisphosphate derivatives 9, 20, and 28, which were found to be substrates of T4 RNA ligase. These thionucleoside 3',5'-bisphosphates were examined as donors for ligation with m3(2,2,7) G5'pppAmUmA, i.e., the 5'-terminal tetranucleotide fragment of U1 snRNA, The 4-thiouridine 3',5'-bisphosphate derivative 9 was found to serve as the most active substrate of T4 RNA ligase with a reaction efficiency of 96%.

Aberrant Cyclization Affords a C-6 Modified Cyclic Adenosine 5′-Diphosphoribose Analogue with Biological Activity in Jurkat T Cells

“…43−46 During the former process, the sulfur is always protected to avoid unnecessary side reactions during phosphitylation and oxidation of the phosphite. 47 Since the synthesis of 6-thio IMP is hampered by the lack of solubility of 6-thioinosine, protecting the sulfur should further solve the solubility issue encountered. Various protecting groups have been used such as cyanoethyl, which can be removed by treatment with DBU.…”

Selenium Derivatization of Nucleic Acids for X‐Ray Crystal‐Structure and Function Studies

“…Encouraged by our exciting discovery and our investigation on the nucleobase pairing and stacking via the atom-specific mutagenesis and crystallography, we have also synthesized for the first time the 6-Se-deoxyguanosine phosphoramidite and 6-Se-dG-containing oligonucleotides (Scheme 12) [51]. Despite the synthesis of the 2'-deoxy-6-selenoguanosine and its derivatives several decades ago [96] [97], as well as the successful incorporation of the 6-S-G derivatives into nucleic acids [98] [99], the synthesis of nucleic acids containing the 6-Se-guanine remained a challenge. Similar to the 4-Se-thymidine synthesis, the synthesis of the key 6-Se-G intermediate 12 was achieved by the selective activation of the guanosine 6-position by TIBS ( ¼ 2,4,6-triisopropylbenzene-1-sulfonyl) group, followed by the Se substitution with the salt sodium 2-cyanoethyl selenide.…”

A New Odorless Procedure for the Synthesis of 2′-Deoxy-6-Thioguanosine and Its Incorporation into Oligodeoxynucleotides

“…[10,11] The excellent synthesis of 2 -deoxy-6-thioguanosine, the phosphoramidite derivative and its incorporation into the ODN have already been reported. [12][13][14][15][16][17][18] One approach utilized the preformed 2 -deoxy-6-thioguanosine to protect the 6-thiocarbonyl group with the cyanoethyl group [12] [13] or the 2,4-dinitorophenyl group. [17,18] These protecting groups are useful, because they can be removed under mild alkaline conditions.…”

“…For the other approach, the 6-carbonyl group was transformed into a labile leaving group such as a halide or arylsulfonyloxy group for displacement with 2-cyanoethanethiol. [14][15][16] In both approaches, odorous agents, such as 2-cyanoethanethiol or hydrogen sulfide, are used. We now describe a new practical synthesis of the 3 -O-phosphoramidite derivative of 6-S-protected 2 -deoxy-6-thioguanosine by using 2-ethylhexyl 3-mercaptopropionate [19,20] as an odorless and inexpensive reagent and its incorporation into the oligodeoxynucleotides.…”