Biochimie 71 (1989) 793- 803 (~) Socirt6 de Chimie biologique/Elsevier, Paris 793 Hairpin structures in synthetic oligodeoxynucleotides- sequence effects on the duplex-to-hairpin transition Luigi Emilio XODO 1., Giorgio MANZINI l, Franco QUADRIFOGLIO 2, Gijs van der MAREL 3 and J.H. van BOOM 3 1Department of Biochemistry, Biophysics and Macromolecular Chemistry, University of Trieste, 1-34127 Trieste, Italy; 2Institute of Biology, Faculty of Medicine, University of Udine, 1-33100 Udine, Italy; and 3Gorlaeus Laboratories, State University, P.O. Box 9502, 2300 RA Leiden, The Netherlands (Received 19-1-1989, accepted after revision 28-3-1989) Summary-- We have synthesized and examined a number of fully and partly self-complementary palin- dromic oligodeoxynucleotides for their ability to assume in solution a unimolecular hairpin structure. The main results obtained by a combined optical and electrophoresis investigation show that: (i) DNA folding needs not be driven by mismatched base pairings over the dyad; fully self-complementary palin- dromic duplexes, comprising regular (CG)n DNA fragments, possess a considerable intrinsic propensity to make intramolecular base pairings; (ii) The duplex-hairpin interconversion is, in general, a slow pro- cess independent of the length and base composition of the palindrome; (iii) The palindromic sequences energetically least favored to form hairpin structures consist of C:G base pairs around the dyad axis and of T:A blocks in the arms of the inverted repeat; (/V) The base composition of the stem strongly influences the hairpin thermal stability. For instance, the substitution of one C:G with one A:T base pair in the stem helix of d(CG)7 diminishes the stability of the hairpin by 9oC. It is found that the stability of the stem helix, in hairpins of defined sequence and with the same loop length, decrt:ases in the order alt~rn~tina_("("- ~ h ..... PC. -. A ~t~_m~ f~- -~...... :__ A ~,- ly dy ~, ........ - . . . . . ~J l 1 att~lHaull~-t~ l, i.e. ~,~ ill po nucleotides. The thermo na- mic parameters for the hairpin-coil transition are reported. duplex-hairpin interconversion / oligodeoxynucleotides / electrophoresis / melting curves / thermodynamics Introduction It is now well established that DNA is characte- rized by a high degree of polymorphism. Its double-stranded helix is no longer regarded as a static rodlike molecule, but as a chain with consi- derable conformational flexibility which may result in aberrant conformations (Z-DNA) [1, 2] as well as imperfect helices containing hairpins, bulges, loops, and so on. Since the control regions of the genome frequently contain invert- ed repeats, i.e. sequences of dyad symmetry (palindromes), it is likely that some biological processes require structural changes in the DNA. Extrusion of palindromic sequences in cruciforms have been found near replication ori- gins in prokaryotes [3] and in eukaryotic viruses [4-6]. Furthermore, cruciform structures are believed to be implied in transcription termina- tion [7]. Recently it has been reported that pro- teins from human extracts [8] and from rat liver nuclei [9] bind specifically to artificial 4-way junctions and not to linear DNA of the same base composition. Since a cruciform is made up of 2 opposite hairpins, studies on the ability of oligodeoxynucleotid"s to form hairpin struc- tures are important. Over the past few years a number of solution studies have been reported on hairpin formation from oligodeoxynucleo- tides [10-25]. Most of these studies have been *Author to whom correspondence should be addressed.