Covalent split protein fragment–DNA hybrids generated through N-terminus-specific modification of proteins by oligonucleotides?

Organic & Biomolecular Chemistry Pub Date: 2008-04-23 DOI: 10.1039/B720013G

Abstract

Semisynthetic protein–DNA hybrid molecules have recently attracted much attention as valuable tools for bioanalytical chemistry and nanobiotechnology. Here we describe a synthetic method for conjugating oligonucleotides to the N-terminus of recombinant proteins. Our strategy involves the conversion of amine-terminated oligonucleotides to thioester-functionalized oligonucleotides by using a bifunctional reagent bearing an N-hydroxysuccinimide ester and benzyl thioester group, followed by native chemical ligation with proteins containing an N-terminal cysteine. We applied this technique to construct split luciferase fragment–DNA hybrid systems in which the catalytic activity of split luciferase is restored by the re-assembly of each fragment through a specific DNA–protein or DNA–DNA interaction. Split protein fragment–DNA hybrids will offer new opportunities to explore the potential of protein–DNA conjugates for various applications.

Graphical abstract: Covalent split protein fragment–DNA hybrids generated through N-terminus-specific modification of proteins by oligonucleotides
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