Hairpin formation within the human enkephalin enhancer region. 1. Kinetic analysis

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Abstract
The 3'-5' cyclic AMP inducible enhancer region of the human enkephalin gene is located within an imperfect palindrome of 23 base pairs, located from -106 to -84 base pairs upstream of the transcriptional start site. Recent evidence has indicated that hairpin formation within this region may be involved in transcriptional regulation of the human proenkephalin gene. A 23-bp synthetic oligonucleotide of this region has been shown to undergo a reversible conformational change from a duplex to a cruciform structure of two hairpins [McMurray, C.T., Wilson, W.D., & Douglass, J.O. (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 666]. Our current studies explore the kinetics and activation energies of the hairpin to duplex transitions of synthetic oligonucleotides under a variety of conditions. Ultraviolet spectroscopic data collected over a range of pH values, ionic strengths, and temperatures are used to determine the reaction rates and activation energies of the hairpin to duplex reaction. The rate of formation of a duplex from two hairpins is a slow second-order process, dependent on both pH and ionic strength. The return from the hairpin state to the duplex state occurs with a high activation energy of 22-41 kcal/mol strand, depending on the conditions. Pseudo-first-order reaction conditions indicate that one of the hairpins, the AC hairpin, is the rate-limiting reactant. These results suggest a model by which the formation of a cruciform might regulate transcription.

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