ENZYMATIC RECOGNITION OF THE BASE-PAIR BETWEEN ISOCYTIDINE AND ISOGUANOSINE

The ability of various polymerases to catalyze the template-directed f ormation of a base pair between isoguanine (iso-G) and isocytosine (is o-C) in duplex oligonucleotides has been investigated. A new procedure was developed for preparing derivatives of deoxyisoguanosine suitable for incorporation into DNA using an automated DNA synthesizer. T7 RNA polymerase, AMV reverse transcriptase, and the Klenow fragment of DNA polymerase all incorporated iso-G opposite iso-C in a template. T4 DN A polymerase did not. Several polymerases also incorporated iso-G oppo site T, presumably through pairing with a minor tautomeric form of iso -G complementary to T. In a template, iso-G directs the incorporation of both iso-C and T when Klenow fragment is the catalyst and only U wh en T7 RNA polymerase is the catalyst. Further, derivatives of iso-C we re found to undergo significant amounts of deamination under alkaline conditions used for base deprotection after automated oligonucleotide synthesis. Both the deamination reaction of iso-C and the ambivalent t automeric forms of iso-G make it unlikely that the (iso-C).(iso-G) bas e pair was a part of information storage molecules also containing the A.T and G.C base pairs found in primitive forms of life that emerged on planet earth several billion years ago. Nevertheless, the extra let ters in the genetic alphabet can serve useful roles in a contemporary laboratory setting.