KAT Ligation for Rapid and Facile Covalent Attachment of Biomolecules to Surfaces

The efficient and bioorthogonal chemical ligation reaction between potassium acyltrifluoroborates (KATs) and hydroxylamines (HAs) was used for the surface functionalizationof a self-assembled monolayer (SAM) with biomolecules. An alkane thioether molecule with one terminal KAT group (S-KAT) was synthesized and adsorbed onto a gold surface, placing a KATgroup on the top of the monolayer (KAT-SAM). As an initial test case, an aqueous solution of a hydroxylamine (HA) derivative of PEG (HA-PEG) was added to this KAT-SAM at room temperature to perform the surface KAT ligation. Quartz crystal microbalance with dissipation (QCM-D) monitoring confirmed the rapid attachment of the PEG moiety onto the SAM.The covalent conjugation of PEG by amide-bond formation was established by complementary surface characterization methods including contact angle, ellipsometry, and X-ray photoelectron spectroscopy (XPS). To test the applicability of this surface KAT ligation for the attachment of biomolecules to the surfaces, this KAT-SAM was subjected to the reaction with HA derivative of protein. A HA-derivatized green fluorescent protein (HA-GFP) was added in dilute concentrations to the KAT-SAM under aqueous conditions and rapid protein attachment was observed in real-time by QCM. Despite the fact that such biomolecules have a variety of unprotected functional groups within their structures, the surface KAT reaction proceeded efficiently in a selective manner. Our results clearly demonstrate the versatile applicability of the KAT ligation for the covalent attachment of a variety of biomolecules onto surfaces under dilute and biocompatible conditions to form stable, natural amide bonds.