ABSTRACT
Poly(2-methylthio-7-deazainosinic acid) [poly(ms2c7I)] was enzymatically synthesized by polymerization of 2-methylthio-7-deazainosine 5'-diphosphate with polynucleotide phosphorylase from Micrococcus luteus in high yield. The homopolymer shows much higher thermal stability than its parent polynucleotides poly(7-deazainosinic acid) [poly(c7I)] and poly(I). Its sigmoidal melting curve and pronounced hypochromicity imply a rigid, ordered structure. Poly(ms2c7I), like poly(2-methylthio-inosinic acid) [poly(ms2I)], does not form a complex with poly(C) because of the bulky 2-methylthio substituent. On the other hand, two poly(ms2c7I) strands form very rigid triple strands with poly(A). Different from poly(I) and poly(c7I) the homopolymer poly(ms2c7I) is very stable against cleavage by nuclease S1 and ribonuclease T2 as expected from its rigid secondary structure.
Subject(s)
Poly I/chemical synthesis , Polyribonucleotides/chemical synthesis , Circular Dichroism , Drug Stability , Indicators and Reagents , Methylation , Micrococcus/enzymology , Nucleic Acid Conformation , Nucleic Acid Denaturation , Polyribonucleotide Nucleotidyltransferase , Spectrophotometry, UltravioletABSTRACT
Poly(8-oxyinosinic acid) (poly O8I), was synthesized by polymerizing 8-oxyinosine diphosphate using the enzyme polynucleotide phosphorylase (PNPase). The polymer formed a 1 : 1 hybrid with polycytidylic acid (poly C). The hybrid was found to induce the production of interferon in the brain of white albino mice and protected mice against Wesselsbron virus (H 10964).
Subject(s)
Poly I/chemical synthesis , Polyribonucleotide Nucleotidyltransferase/metabolism , Polyribonucleotides/chemical synthesis , Animals , Chemical Phenomena , Chemistry, Physical , Dose-Response Relationship, Drug , Hydrogen-Ion Concentration , Interferon Inducers , Lethal Dose 50 , Mice , Poly I/pharmacology , Spectrophotometry, UltravioletSubject(s)
Interferon Inducers , Polyribonucleotides/chemical synthesis , RNA, Double-Stranded/chemical synthesis , Micrococcus/enzymology , Poly I/chemical synthesis , Poly I/isolation & purification , Polyribonucleotide Nucleotidyltransferase , Polyribonucleotides/pharmacology , RNA, Double-Stranded/pharmacologyABSTRACT
Poly (2'-chloro-2'-deoxyinosinic acid) [poly(Icl)] was synthesized from Icl 5'-DP by polymerization with polynucleotide phosphorylase. UV absorption properties of poly(Icl) are very similar to those of poly(I). Poly(Icl) adopted a multi-stranded ordered form in the presence of 0.95M Na ion. The Tm value of this form was 36 degrees, which resembles that of poly(I) quadruple-stranded form at high salt. CD spectra also suggested presence of these two forms. Upon mixing with poly(C), poly-(Icl) forms a double-stranded 1 : 1 complex, which had very similar Tm-log[Na+] relationship to that of poly(I) . poly(C). Thus it was concluded that the chlorine substitution at 2'-position of the polynucleotide had the similar effect to OH on physical properties of polynucleotides.
Subject(s)
Poly I , Polyribonucleotides , Circular Dichroism , Drug Stability , Hot Temperature , Nucleic Acid Conformation , Poly C , Poly I/chemical synthesis , Polyribonucleotide Nucleotidyltransferase , Polyribonucleotides/chemical synthesis , Spectrophotometry, UltravioletABSTRACT
Chemically synthesized 2-azaadenosine 5'-diphosphate (n2ADP) and 2-azainosine 5'-diphosphate (n2IDP) were polymerized to yield poly(2-azaadenylic acid), poly(n2A), and poly(2-azainosinic acid), poly(n2I), using Escherichia coli polynucleotide phosphorylase. In neutral solution, poly(n2A) and poly(n2I) had hypochromicities of 32 and 5.5%, respectively. Poly(n2A) formed an ordered structure, which had a melting temperature (Rm) of 20 degrees C at 0.15 M salt concentration. Upon mixing with poly(U), poly(n2A) formed a 1 : 2 complex with Tm of 41 degrees C at 0.15 M salt concentration. Poly(n2A) and poly(n2I) formed three-stranded complexes with poly(I), and poly(A), respectively. Poly(n2A) . 2poly(I), poly(A) . 2poly(n2I), and poly(n2A) . 2poly(n2I) complexes had Tm values of 23, 48, and 31 degrees C at 0.15 M salt concentration, respectively. Poly(n2I) formed a double-stranded complex with poly(C), but its Tm was very low.
Subject(s)
Poly A , Poly I , Polyribonucleotides , Circular Dichroism , Escherichia coli/enzymology , Nucleic Acid Denaturation , Poly A/chemical synthesis , Poly I/chemical synthesis , Polyribonucleotide Nucleotidyltransferase , Polyribonucleotides/chemical synthesis , Spectrophotometry, UltravioletABSTRACT
Poly (2'-deoxy-2'-fluoroinosinic acid) [ poly(If)] was synthesized by polymerization of 2'-deoxy-2'-fluoroinosine 5'-diphosphate catalyzed by Escherichia coli polynucleotide phosphorylase. Although the UV absorption properties of poly(If) closely resembled those of poly(I), thermal melting curves at Na+ concentrations of 0.15M and 0.75M suggested two ordered structures for poly(If) neutral form. CD psectra taken at 0.15M Na+ concentration showed rather larger amplitudes in both a peak at 273 nm and a trough at 246 nm, suggesting rather strong vertical stacking of bases. When complexed with poly(C), poly(If) forms a double-stranded complex, poly(If).poly(C) which has Tm's higher by 10-20 degrees than those of poly(If).poly(C) measured under the same conditions. The CD spectrum of this complex resembled that of poly(I).poly(C). The effect of the fluorine atom at the 2'-position on thermal stability of polynucleotides is discussed.
Subject(s)
Poly I/chemical synthesis , Polyribonucleotides/chemical synthesis , Circular Dichroism , Hot Temperature , Nucleic Acid Conformation , Nucleic Acid Denaturation , Poly I-C , Polydeoxyribonucleotides/chemical synthesis , Spectrophotometry, UltravioletABSTRACT
Poly 7-deazainosinic acid has been prepared by the deamination and phosphorylation of tubercidin and the nucleoside diphosphate was polymerised using polynucleotide phosphorylase. The polymer has similar physical properties to poly(I), but has a low thermal stability in the double-stranded complex with poly(C). Poly(7-deaza I), in contrast, forms a more stable triple-stranded complex with poly(A) than 2 poly(I). poly(A), presumably due to the higher pK value.
