Synthesis of 2'-deoxy-2'-fluoroguanyl-(3',5')-guanosine

The protected analogue of 2-amnio-6-chloropurine arabinoside (3b) was subjected to reaction with diethylaminosulfur trifluoride (DAST) and subsequently treated with NaOAc in Ac2O/AcOH to give N2, O3', O5'-triacetyl-2'-deoxy-2'-fluoroguanosine (5a). After deacetylation of the sugar moiety and protection of 5'-OH by a 4,4'-dimethoxytrityl group, this nucleoside component was converted to 2'-deoxy-2'-fluoroguanyl-(3',5')-guanosine (6c, GfpG).     

Analogs of adenosine 3',5'-cyclic phosphate. II. Synthesis and enzymatic activity of derivatives of 1,N6-ethenoadenosine 3',5'-cyclic phosphate

The reactions of chloroacetaldehyde with adenosine 3′,5′-cyclic phosphate, and with several analogs modified at C₈ of the purine ring or C₅, of the sugar, lead to the corresponding 1,N⁶-etheno derivatives^d. Similar reactions using other 2-bromoaldehydes or phenacyl bromide give 1,N⁶-ethenonucleotides substituted at the α- or β-positions of the etheno bridge respectively. The ability of these compounds to activate the protein kinases from rabbit muscle and calf brain has been evaluated over a wide range of concentrations. While no derivative proved to be more active than adenosine 3′,5′-cyclic phosphate itself using the enzyme from rabbit muscle, a wide spectrum of activities was found using that from calf brain.     

Inhibition of cGMP-dependent protein kinase by (Rp)-guanosine 3',5'-monophosphorothioates

The activation of the cGMP-dependent protein kinase and cAMP-dependent protein kinase by the diastereomers of guanosine 3',5'-monophosphorothioate, (Sp)-cGMPS and (Rp)-cGMPS, and 8-chloroguanosine 3',5'-monophosphorothioate, (Sp)-8-Cl-cGMPS and (Rp)-8-Cl-cGMPS, was investigated using the peptide Kemptide as substrate. The (Sp)-diastereomers, which have an axial exocyclic sulfur atom, bound to the cGMP-dependent protein kinase and stimulated its phosphotransferase activity. In contrast, the (Rp)-isomers, which have an equatorial exocyclic sulfur atom, bound to the enzyme without stimulation of its activity. (Rp)-cGMPS and (Rp)-8-Cl-cGMPS antagonized the activation of the cGMP-dependent protein kinase with a Ki of 20 microM and 1.5 microM, respectively. (Rp)-cGMPS also antagonized the activation of cAMP-dependent protein kinase with a Ki of 20 microM. In contrast, (Rp)-8-cGMPS ws a weak inhibitor of the cAMP-dependent protein kinase with a Ki of 100 microM. (Rp)-8-Cl-cGMPS appears to be a rather selective inhibitor of the cGMP-dependent protein kinase and may be a useful tool for studying the role of cGMP in broken and intact cell systems.     

Synthesis and enzymatic deprotection of biodegradably protected dinucleoside-2',5'-monophosphates: 3-(acetyloxy)-2,2-bis(ethoxycarbonyl)propyl phosphoesters of 3'-O-(acyloxymethyl)adenylyl-2',5'-adenosines

As a first step towards a viable prodrug strategy for short oligoribonucleotides, such as 2–5A and its congeners, adenylyl‐2′,5′‐adenosines bearing a 3‐(acetyloxy)‐2,2‐bis(ethoxycarbonyl)propyl group at the phosphate moiety, and an (acetyloxy)methyl‐ or a (pivaloyloxy)methyl‐protected 3′‐OH group of the 2′‐linked nucleoside have been prepared. The enzyme‐triggered removal of these protecting groups by hog liver carboxyesterase at pH 7.5 and 37° has been studied. The (acetyloxy)methyl group turned out to be too labile for the 3′‐O‐protection, being removed faster than the phosphate‐protecting group, which results in 2′,5′‐ to 3′,5′‐isomerization of the internucleosidic phosphoester linkage. In addition, the starting material was unexpectedly converted to the 5′‐O‐acetylated derivative. (Pivaloyloxy)methyl group appears more appropriate for the purpose. The fully deprotected 2′,5′‐ApA was accumulated as a main product, although, even in this case, the isomerization of the starting material takes place.     

Synthesis and conformational properties of O-beta-D-ribofuranosyl-(1'-2')-guanosine and (adenosine)-5'-phosphate

The efficient synthesis of Grp and Arp, minor tRNA components, has been developed.     

Activity of tubercidin-, toyocomycin-, and sangivamycin-3',5'-cyclic phosphates and related compounds with some enzymes of adenosine-3',5'-cyclic phosphate metabolism

Rat liver nuclei contain at least two DNA polymerases that can be separated by extracting the nuclei with 5% Brij 58. The loosely-bound activity increases little or not at all after partial hepatectomy and is insensitive to cytosine arabinoside triphosphate (araCTP). The tightly-bound enzyme activity rises along with DNA replication and is inhibited by araCTP.     

Theoretical conformation analysis of cytidine 2',3'-cyclic phosphate]

On the basis of calculations of conformational states of cytidine 2',3'-cyclic phosphate it is shown that the syn-form with a gauche-trans position of the exocyclic group C(5')H2OH is preferential.     

Diphtheria toxin can simultaneously bind to its receptor and adenylyl-(3',5')-uridine 3'-monophosphate

Diphtheria toxin that was bound to receptors on BS-C-1 cells was able to bind approximately 1 molar equiv of adenylyl-(3',5')-uridine 3'-monophosphate (ApUp). In contrast, receptor-bound CRM197, a mutant form of toxin with greatly diminished affinity for dinucleotides, did not bind ApUp. Affinity of the dinucleotide for receptor-bound toxin differed from that for free toxin by less than an order of magnitude. These results indicate that the receptor site and the ApUp site on the toxin do not significantly overlap.