Phosphorothioate analogues of 2',5'-oligoadenylate. Enzymatic synthesis, properties, and biological activities of 2',5'-phosphorothioates from adenosine 5'-O-(2-thiotriphosphate) and adenosine 5'-O-(3-thiotriphosphate).
The chiral and achiral phosphorothioate analogues of 2',5'-oligoadenylates (2-5A) have been enzymatically synthesized from the Sp and Rp isomers of adenosine 5'-O-(2-thiotriphosphate) [(Sp)-ATP beta S and (Rp)-ATP beta S, respectively] and adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S) by 2-5A synthetase from L929 cells and lysed rabbit reticulocytes. These 2',5'-phosphorothioate analogues were separated, purified, and structurally characterized. While ATP gamma S and (Sp)-ATP beta S were as efficient substrates for the 2-5A synthetase as was ATP, (Rp)-ATP beta S was more than 50-fold less efficient a substrate. The beta- and gamma-phosphorothioates were more resistant to enzymatic hydrolysis than was authentic 2-5A. Compared to 2-5A, there were marked differences in the biological activities of the 2',5'-phosphorothioates as determined by (i) binding to 2-5A-dependent endoribonuclease (RNase L), (ii) activation of RNase L to hydrolyze RNA, and (iii) inhibition of protein synthesis in intact L929 cells. These studies extend previous reports on the elucidation of the stereochemical requirements of 2-5A synthetase and RNase L [Karikó, K., Sobol, R. W., Jr., Suhadolnik, L., Li, S. W., Reichenbach, N. L., Suhadolnik, R. J., Charubala, R., & Pfleiderer, W. (1987) Biochemistry (first of three papers in this issue); Karikó, K., Li, S. W., Sobol, R. W., Jr., Suhadolnik, R. J., Charubala, R., & Pfleiderer, W. (1987) Biochemistry (second of three papers in this issue)] with the phosphorothioate analogues of 2-5A.(ABSTRACT TRUNCATED AT 250 WORDS)
Department of Biochemistry, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.
R J
RJ
Suhadolnik
C
C
Lee
K
K
Karikó
S W
SW
Li
D000227:Adenine Nucleotides / Q000138:chemical synthesis* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives*; D000818:Animals; D004355:Drug Stability; D004722:Endoribonucleases / Q000378:metabolism*; D007700:Kinetics; D007739:L Cells / Q000201:enzymology; D051379:Mice; D009843:Oligoribonucleotides / Q000138:chemical synthesis* / Q000378:metabolism; D012335:RNA, Ribosomal / Q000378:metabolism; D013329:Structure-Activity Relationship; D013873:Thionucleotides / Q000138:chemical synthesis* / Q000378:metabolism
D000227:Adenine Nucleotides; D009843:Oligoribonucleotides; D012335:RNA, Ribosomal; D013873:Thionucleotides; C022571:adenosine 5'-O-(3-thiotriphosphate); C022570:adenosine 5'-O-(2-thiotriphosphate); C023505:2',5'-oligoadenylate; D000255:Adenosine Triphosphate; D004722:Endoribonucleases; C031956:2-5A-dependent ribonuclease