Efforts toward expansion of the genetic alphabet: Optimization of interbase hydrophobic interactions

Six novel unnatural nucleobases have been characterized that form stable ba se pairs in duplex DNA, relying nor on hydrogen bonds, but rather on interb ase hydrophobic interactions. These nucleobases are derivatives of the hydr ophobic base pair between 7-azaindole (7AI) and isocarbostyril (ICS). Deriv atives of 7AI and ICS were examined that have increased hydrophobic surface area, as well as increased polarizability. As observed with 7AI and ICS, t hese derivatives are recognized as substrates by Klenow fragment of Escheri chia coli DNA polymerase I. The unnatural base pair between pyrrolopyrizine (PP) and C3-methylisocarbostyril (MICS) is enzymatically incorporated into DNA with high efficiency (k(cat)/K-M = 10(6) M-1 min(-1)) and moderate sel ectivity. These studies represent a significant step toward the generation of astable, orthogonal base pair that can be enzymatically incorporated int o DNA with good fidelity.