Oligomerization of Adenosin-5 0 -O-ylmethylphosphonate, an Isopolar AMP Analogue: Evaluation of the Route to Short Oligoadenylates Martina Pressova ´, 1 Milos ˇ Bude ˇs ˇı ´nsky ´, 1 Ivana Kos ˇiova ´, 1 Vladimı´r Kopecky ´ Jr., 2 Josef Cvac ˇka, 1 Va ´clav Kas ˇic ˇka, 1 Ondr ˇej S ˇ ima ´k, 1 Zdene ˇk Toc ˇı ´k, 1 Ivan Rosenberg 1 1 Institute of Organic Chemistry and Biochemistry, Academy of Sciences, v. v. i., Flemingovo 2, 16610 Prague 6, Czech Republic 2 Institute of Physics, Faculty of Mathematics and Physics, Charles University, Ke Karlovu 5, 12116 Prague 2, Czech Republic Received 28 July 2009; revised 13 October 2009; accepted 13 October 2009 Published online 20 October 2009 in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/bip.21329 This article was originally published online as an accepted preprint. The ‘‘Published Online’’ date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley. com INTRODUCTION T he short adenine-based 5 0 -phosphorylated 2 0 ,5 0 -oli- goribonucleotides (trimers, tetramers, and higher ones) have been found to be natural activators of RNase L, the enzyme playing a crucial role in the cel- lular antiviral defense mechanism. 1 2 0 ,5 0 -Oligoadeny- lates acting at subnanomolar concentrations cause the dime- rization of RNase L subunits into an active form capable of cleaving ssRNA. 2 Structure-activity studies on the structur- ally diverse, modified oligoadenylates, and related com- pounds made within last two decades provided basic knowl- edge on the structural requirements of the enzyme for partic- ular parts of the oligoadenylate molecule. 1 Oligomerization of Adenosin-5 0 -O-ylmethylphosphonate, an Isopolar AMP Analogue: Evaluation of the Route to Short Oligoadenylates Correspondence to: M. Bude ˇs ˇı ´nsky ´ ; e-mail: budesinsky@uochb.cas.cz or Zdene ˇk Toc ˇı ´k; e-mail: tocik@uochb.cas.cz or I. Rosenberg; e-mail: ivan@uochb.cas.cz ABSTRACT: In an attempt to prepare a library of short oligoadenylate analogues featuring both the enzyme-stable internucleotide linkage and the 5 0 -O-methylphosphonate moiety and thus obtain a pool of potential RNase L agonists/antagonists, we studied the spontaneous polycondensation of the adenosin-5 0 -O- ylmethylphosphonic acid (p c A), an isopolar AMP analogue, and its imidazolide derivatives employing N,N 0 -dicyclohexylcarbodiimide under nonaqueous conditions and uranyl ions under aqueous conditions, respectively. The RP LC–MS analyses of the reaction mixtures per se, and those obtained after the periodate treatment, along with analyses and separations by capillary zone electrophoresis, allowed us to characterize major linear and cyclic oligoadenylates obtained. The structure of selected compounds was supported, after their isolation, by NMR spectroscopy. Ab initio calculation of the model structures simulating the AMP- imidazolide and p c A-imidazolide offered the explanation why the latter compound exerted, in contrast to AMP- imidazolide, a very low stability in aqueous solutions. # 2009 Wiley Periodicals, Inc. Biopolymers 93: 277–289, 2010. Keywords: spontaneous oligomerization; oligoadenylates; phosphonate internucleotide linkage; imidazolides; uranyl ions catalysis Contract grant sponsor: EU Contract grant number: EMIL-FW6 503569 Contract grant sponsor: Ministry of Education, CR Contract grant numbers: LC06077, LC06061 Contract grant sponsor: Ministry of Education, CR Contract grant number: MSM 0021620835 Contract grant sponsor: Academy of Sciences CR Contract grant number: KAN200520801 Contract grant sponsor: Czech Science Foundation Contract grant numbers: 203/09/0820, 203/09/0193 Contract grant sponsor: Institute Research Project Contract grant number: Z40550506 V V C 2009 Wiley Periodicals, Inc. Biopolymers Volume 93 / Number 3 277