Elasticity of the transition state for oligonucleotide hybridization

Kevin D. Whitley, Matthew J. Comstock, Yann R. Chemla

Research output: Contribution to journalArticle

Abstract

Despite its fundamental importance in cellular processes and abundant use in biotechnology, we lack a detailed understanding of the kinetics of nucleic acid hybridization. In particular, the identity of the transition state, which determines the kinetics of the twostate reaction, remains poorly characterized. Here, we used optical tweezers with single-molecule fluorescence to observe directly the binding and unbinding of short oligonucleotides (7-12 nt) to a complementary strand held under constant force. Binding and unbinding rate constants measured across a wide range of forces (1.5-20 pN) deviate from the exponential force dependence expected from Bell's equation. Using a generalized force dependence model, we determined the elastic behavior of the transition state, which we find to be similar to that of the pure single-stranded state. Our results indicate that the transition state for hybridization is visited before the strands form any significant amount of native base pairs. Such a transition state supports a model in which the rate-limiting step of the hybridization reaction is the alignment of the two strands prior to base pairing.

Original languageEnglish (US)
Pages (from-to)547-555
Number of pages9
JournalNucleic acids research
Volume45
Issue number2
DOIs
StatePublished - Jan 1 2017

ASJC Scopus subject areas

  • Genetics

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