MECHANISM OF RIBOSYLATION OF ADENINE

Transcription

1 OC8 Ribosylation of Adenine 177 MECHANISM OF RIBOSYLATION OF ADENINE Jerzy BORYSKI* and Grzegorz FRAMSKI Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, PL-61704, Poznań, Poland; Depending on reaction conditions and the presence of N 6 -protecting groups, ribosylation of adenine proceeds via different kinetic products, which finally undergo an intermolecular transglycosylation to adenosine. While initial 3-ribosylation takes place in strongly acidic media, the 7-regioisomer of adenosine is a kinetic product in ribosylation of unprotected adenine by using the silyl approach. Unexpectedly, the silyl method in the case of N 6 -acyladenine derivatives gives a new kinetic product of ribosylation, 1-(β-D-ribo- furanosyl)adenine. INTRODUCTION Acid-catalyzed ribosylation of heterocyclic bases, one of the most important reactions in the nucleoside chemistry, represents a complex chemical reaction. The process is initiated by the generation of a reactive sugar cation from an appropriate sugar derivative (e.g. peracylated ribose or its 1-halogenated derivative). The cation reacts then with a heterocyclic base. However, the sugar residue cannot be attached directly to its ulimate position (e.g. N9 in majority of purine bases), because that position is already blocked by proton, in the most stable tautomeric form of the base. Therefore, the sugar may be attached to any alternative position, i.e. N3 or N7 in the purine series. This results in the formation of a kinetic product, which then undergoes an intermolecular rearrangement (transglycosylation) to a more stable regioisomer. In this way the structure of thermodynamic product of ribosylation corresponds to that of the most stable tautomer of a starting heterocyclic base. In the case of adenine, ribosylation performed under strongly acidic conditions (30% HBr in acetic acid) gave a mixture of adenosine and its regioisomer, 3-(β-D-ribofuranosyl)adenine (3-isoadenosine) 1. In line with that observation, the first proposed mechanism 2 postulates the following sequence of events: (i) initial ribosylation at N3, (ii) second ribosylation at N9 with the formation of 3,9-bis-ribosyladenine, and (iii) its decomposition to adenosine. 3-Riboadenine was obtained also by applying the mercury procedure 3. Its structure and kinetic character have been fully confirmed 4,5. In 1970th that mechanism was extended for ribosylation reactions of all purine bases.

2 178 Boryski, Framski: More recently, however, it has been shown that only N7 and N9 atoms can serve as glycosyl donors or acceptors in glycosylation and transglycosylation reactions in the guanine series 6,7. In addition, there are some literature reports on obtaining of 7-(β-D-ribofuranosyl)adenine in the ribosylation of adenine RESULTS AND DISCUSSION In our systematic reinvestigation of mechanisms of the N-glycosylic bond formation, we performed a series of experiments to establish the factors responsible for either 3 9 or7 9 mechanism in the ribosylation of adenine. This time we looked more closely at the silyl method, since at present it is the most common synthetic procedure. Thus, adenine (1) was silylated with hexamethyldisilazane (HMDS) and then subjected to ribosylation with 1,2,3,5-tetra-O-acetyl-β-D-ribofuranose in the presence of trimethylsilyl triflate (TMSTf) (Scheme 1). After 80 min the reaction mixture contained 24% of the 7-isomer (2; isolated by column chromatography), along with the prevailing amounts of triacetyladenosine (3). When heating was continued for a longer time, the 9-regioisomer 3 was the only remaining product. Similarly, the isolated product 2 could be quantitatively isomerized to 3 on refluxing in chlorobenzene in the presence of p-toluenesulfonic acid. This shows clearly that the 7-riboadenine is a kinetic product in the ribosylation of adenine. Most probably, the 7 9 SCHEME 1

3 Ribosylation of Adenine 179 transglycosylation proceeds via a 7,9-diribosyl intermediate, likewise in the guanine series. However, this product was not isolated from the reaction mixture in the present work. The kinetic product 2 was then deprotected with aqueous ammonia in methanol to give 7-(β-D-ribofuranosyl)adenine (4). SCHEME 2 Therefore, we could expect an initial 7-ribosylation in the reaction of N 6 -acylated derivatives of adenine (5; Scheme 2), performed according to the Vorbrüggen s procedure. In our experiments, the compounds of the type 5 (R = benzoyl or isobutyryl) were silylated with N,O-bis-trimethylsilylacetamide (BSA). Surprisingly, their ribosylation with 1,2,3,5-tetra- O-acetyl-β-D-ribofuranose in the presence of TMSTf gave, in addition to the main product 7, a new product of the structure of 1-regioisomer (6). Indeed, that compound was a kinetic product in ribosylation of 5: (i) it underwent isomerization to 7 after a prolonged reaction time; (ii) its reaction with 2-acetoxyethyl acetoxymethyl ether resulted in the formation of acycloadenosine derivative 8. The deprotection of 6 with sodium methoxide gave

4 180 Boryski, Framski: 1-(β-D-ribofuranosyl)adenine (9). Its structure was confirmed by the 1 H, 13 C and 15 N NMR (1D & 2D techniques, NOE), and X-ray analysis. Moreover, a careful chromatographic separation of the reaction mixture allowed us to isolate another minor product of ribosylation of 5 (R = isobutyryl), 1,9-bis-(β-D-ribofuranosyl)adenine derivative (10; yield 5%). Apparently, it is an intermediate in the 1 9 transglycosylation reaction. Unlike quaternary 7,9-bis-ribosylpurine derivatives, intermediates in 7 9 transglycosylation of 6-oxopurine nucleosides 11, compound 10 was quite stable, and its decomposition to 7 required the presence of an acid catalyst. The obtaining of 1-regioisomer of adenosine (6) as a kinetic product of ribosylation is not in line with the previous report of Ryan, Acton and Goodman 8, who have reported the formation of 7-β-D-ribofuranoside in the ribosylation of N 6 -benzoyladenine under similar conditions. That product, however, was not fully characterized, and the structure elucidation of 7-isoadenosine based only on IR and UV data. It is worthy to note that the formation of 1-(β-D-ribofuranosyl)adenine as a possible kinetic intermediate has been anticipated by Vorbrüggen and Höfle 12, but so far this has never been proved experimentally. We thank Prof. Zofia Gdaniec (Institute of Bioorganic Chemistry) for the NMR analysis, and Prof. Maria Gdaniec (Faculty of Chemistry, Adam Mickiewicz University) for the X-ray analysis. REFERENCES 1. Leonard N. J., Laursen R. A.: Biochemistry 1965, 4, Watanabe K. A., Hollenberg D. H., Fox J. J.: J. Carbohydr., Nucleosides, Nucleotides 1974, 1, Shimizu B., Miyaki M.: Chem. Pharm. Bull. 1970, 18, Shimizu B., Miyaki M.: Chem. Pharm. Bull. 1970, 18, 1446.

5 Ribosylation of Adenine Framski G., Manikowski A., Zandecki T., Boryski J.: Nucleic Acids Res. 2003, Supplement No. 3, Boryski J.: Nucleosides Nucleotides 1996, 15, 771; and references therein. 7. Boryski J.: J. Chem. Soc., Perkin Trans , Ryan K. J., Acton E. M., Goodman L.: J. Org. Chem. 1971, 36, Nakazaki N., Sekiya M., Yoshino T., Ishido Y.: Bull. Chem. Soc. Jpn. 1973, 46, Itoh T., Mizuno Y.: Heterocycles 1976, 5, Boryski J., Manikowski A.: Nucleosides Nucleotides 1999, 18, Vorbrüggen H., Höfle G.: Chem. Ber. 1981, 114, 1256.