Pegylation of RNA Spiegelmers by a Novel Widely Applicable Two-Step Process for the Conjugation of Carboxylic Acids to Amino-Modified Oligonucleotides

摘要

The reaction between N-hydroxy succinimide (NHS) ester-activated carboxylic acids and amino-modified nucleic acids is commonly used for the postsynthetic modification of oligonucleotides. Here, we report a two-step variation of the method in which theNHS ester is replaced by the corresponding parent carboxylic acid. In the first step, the carboxylic acid is activated with a standard peptide coupling reagent like HBTU in an anhydrous water-miscible aprotic organic solvent. In the second step, the solution of the activated carboxylic acid is added to the amino-modified oligonucleotide in water. The method is demonstrated using 40-kDa polyethylene glycol (PEG) carboxylic acid and biotin as examples. Recycling of the carboxylic acid, which is typicallyused in molar excess over the nucleic acid, is shown for the conjugation with 40-kDa PEG carboxylic acid. This conjugation method is generally applicable to the conjugation of carboxylic acids to amino-modified oligonucleotides, thus enabling the attachment of small to large molecular entities such as dyes, tags, peptides, and other macromolecules.