The Parallel and Antiparallel Triplex Formation and Stability of Self Complementary Oligonucleotides Containing 2′-Fluoro-arabinosyl Thymine and 5-Methyl-2′-Deoxycytidine

Abstract

We examined the effects of 1–(2-deoxy -2-fluoro-β-D-arabinofuranosyl)-thymine (or FMAU, a potent antiviral nucleoside) on the stability of duplex and triplexes. When compared the stability of the self-complementary 5′-A₅T₅ duplex with 5′-A₅X₅ (X = FMAU), duplex containing FMAU has much higher melting temperature (T_m). 5′-A₆T₅T₃X₃T₅F₃X₃ and T₃X₃T₅A₆T₅F₃X₃ form the parallel and antiparallel triplexes T₃X₃: A₆:X₃X₃, respectively. The former exhibited the typical T:A:T triplex behavior with only one melting temperature at 70 °C and 45 °c in 1.0 M and 0.2 M NaCl solution, respectively, whereas the latter has two T_m values at 56 °C and 28 °C in 1.0 M solution. FMAU clearly stabilize the triplex structure as A₆T₂₂ which forms the parallel triplex T₆:A₆:T₆ has also only one T_m at 54 °C and 37 °C in high and iow salt concentration solutions, respectively. A 31mer 5′-TCCTCCTTTTTTAGGAGGATTTTTTGGTGGT and 5′-TCCTCCTTTTTTAGGAGGATTTTTTX'X'TX'X'T (X' = 2′-deoxy-5-methylcytidine) were prepared to study their triplex forming potential. The former was found to have a week interaction of the Watson-Crick duplex with the mismatched third-strand at all pH. The latter formed a stable triplex at lower pH consistent with required protonation on the 5-methylcytosine base. For these studies we developed a simple PC desktop spreadsheet program to calculate the first derivative profile of the melting curve data.

This paper is dedicated to Prof. Jacques H. van Boom on the occasion of his 60th birthday.