Facile Functionalization of Colloidal Gold Nanorods by the Specific Binding of an Engineered Protein that Is Preferred over CTAB Bilayers

The functionalization of gold nanorods (GNRs) is an important step in the exploitation of their unique optical properties for chemical and biological applications such as surface-enhanced Raman spectroscopy (SERS), chemical and biological sensing, bioimaging, and photothermal therapy. In the highyield synthesis of GNRs, cetyltrimethylammonium bromide (CTAB) directs the anisotropic growth of nanogold seeds and necessarily exists as a bilayer on the surface of GNRs to stabilize the GNRs. The absence of functional moieties in CTAB requires functionalization steps for conjugation, and poor stability of the CTAB bilayer also limits the use of GNRs necessitating complementary materials and processes for further application. Replacement of the CTAB bilayer with small, thiolated compounds or other macromolecules is typically performed to provide functional groups to enhance colloidal stability and functionality of GNRs. In other approaches, direct coating on GNRs using a variety of methods that involve coating with silica, polyelectrolytes, biopolymers, and phospholipids have been studied. Despite of the diversity of functionalization materials, the most common method for the attachment of functional materials is accomplished through classic gold–thiol bond chemistry or electrostatic interactions. In addition, functionality of the materials has mainly focused on providing chemical groups such as amino or carboxylic groups, except in few cases that involve biotin. Herein, we demonstrate a conceptually different method to functionalize GNRs. Our new approach is based on a bifunctional macromolecule with gold-specific binding affinity. In our earlier study, we developed an engineered fusion protein in which gold-binding polypeptide (GBP) was fused with Staphylococcal Protein A (SpA) to recognize the non-antigenic (Fc) portion of antibodies. The recombinant fusion protein, GBP– SpA, showed specific adsorption on different types of gold nanostructures. A facile functionalization method of GNRs using GBP-SpA is described here, which extends to a biomolecular sensing application. As the functionalization of colloidal GNRs has become more challenging owing to the dense capping of CTAB and other aggregation problems, we first examined the interactions among GNRs, the CTAB bilayer, and GBP– SpA to design a functionalization method. Significantly improving stability by the in situ removal of free CTAB, we functionalized GNRs to this GNR–GBP–SpA hybrid nanomaterial through specific interactions between the gold surface and the GBP domain, as illustrated in Scheme 1.