Kawamura K, Ferris J P
Department of Chemistry, Rensselaer Polytechnic Institute, Troy, New York 12180-3590, USA.
J Am Chem Soc. 1994;116(17):7564-72. doi: 10.1021/ja00096a013.
The rate constants for the condensation reaction of the 5'-phosphorimidazolide of adenosine (ImpA) to form dinucleotides and oligonucleotides have been measured in the presence of Na(+)-volclay (a Na(+)-montmorillonite) in pH 8 aqueous solution at 25 degrees C. The rates of the reaction of ImpA with an excess of adenosine 5'-monophosphoramidate (NH2pA), P1,P2-diadenosine 5',5'-pyrophosphate (A5'ppA), or adenosine 5'-monophosphate (5'-AMP or pA) in the presence of the montmorillonite to form NH2pA3'pA, A5'ppA3'pA, and pA3'pA, respectively, were measured. Only 3',5'-linked products were observed. The magnitude of the rate constants decrease in the order NH2pA3'pA > A5'-ppA3'pA > pA3'pA. The binding of ImpA to montmorillonite was measured, and the adsorption isotherm was determined. The binding of ImpA to montmorillonite and the formation of higher oligonucleotides is not observed in the absence of salts. Mg2+ enhances binding and oligonucleotide formation more than Ca2+ and Na+. The rate constants for the oligonucleotide formation were determined from the reaction products formed from 10 to 40 mM ImpA in the presence of Na(+)-montmorillonite using the computer program SIMFIT. The magnitudes of the rate constants for the formation of oligonucleotides increased in the order 2-mer < 3-mer < 4-mer ... 7-mer. The rate constants for dinucleotide and trinucleotide formation are more than 1000 times larger than those measured in the absence of montmorillonite. The rate constants for the formation of dinucleotide, trinucleotide, and tetranucleotide are 41,2.6, and 3.7 times larger than those for the formation of oligo(G)s with a poly(C) template. The hydrolysis of ImpA was accelerated 35 times in the presence of the montmorillonite. The catalytic ability of montmorillonite to form dinucleotides and oligonucleotides is quantitatively evaluated and possible pathways for oligo(A) formation are proposed.
在25℃的pH 8水溶液中，于存在Na(+)-蒙脱石（一种钠蒙脱石）的情况下，测定了腺苷5'-磷酰咪唑酯（ImpA）缩合形成二核苷酸和寡核苷酸的反应速率常数。测定了在蒙脱石存在下，ImpA与过量的腺苷5'-单磷酸酰胺（NH2pA）、P1,P2-二腺苷5',5'-焦磷酸（A5'ppA）或腺苷5'-单磷酸（5'-AMP或pA）反应分别形成NH2pA3'pA、A5'ppA3'pA和pA3'pA的速率。仅观察到3',5'-连接的产物。速率常数的大小顺序为NH2pA3'pA > A5'-ppA3'pA > pA3'pA。测定了ImpA与蒙脱石的结合，并确定了吸附等温线。在无盐的情况下未观察到ImpA与蒙脱石的结合及更高寡核苷酸的形成。Mg2+比Ca2+和Na+更能增强结合和寡核苷酸的形成。使用计算机程序SIMFIT，根据在Na(+)-蒙脱石存在下由10至40 mM ImpA形成的反应产物，测定了寡核苷酸形成的速率常数。寡核苷酸形成的速率常数大小顺序为2聚体 < 3聚体 < 4聚体……7聚体。二核苷酸和三核苷酸形成的速率常数比在无蒙脱石情况下测定的速率常数大1000倍以上。二核苷酸、三核苷酸和四核苷酸形成的速率常数分别比用聚（C）模板形成寡聚（G）的速率常数大41、2.6和3.7倍。在蒙脱石存在下，ImpA的水解加速了35倍。定量评估了蒙脱石形成二核苷酸和寡核苷酸的催化能力，并提出了寡聚（A）形成的可能途径。
