5′-Amino Pyrene Provides a Sensitive, Nonperturbing Fluorescent Probe of RNA Secondary and Tertiary Structure Formation

Blocked phosphoramidites of 5′-amino-5′-deoxycytidine and 5′-amino-5′-deoxyuridine have been prepared and coupled to the 5′-end of oligoribonucleotides. These 5′-amino RNA oligomers may be readily conjugated to a variety of commercially available amine reactive fluorescent dyes, including succinimidyl ester derivatives. The synthesis, purification, and properties of 5′-pyrene-labeled RNA oligomers are described. These oligomers are shown to be unusually attractive probes for studies of oligomer-oligomer and oligomer-ribozyme interactions. In particular, 5′-pyrene oligomers are stable and very sensitive to environment while having minimal perturbations on the thermodynamics of secondary and tertiary structure formation in RNA. Thus the 5′-positioning of pyrene with a short linker appears to provide an ideal probe for the binding and dynamics of RNA substrates.