Bioorthogonal Modification of the Major Sheath Protein of Bacteriophage M13: Extending the Versatility of Bionanomaterial Scaffolds

Abstract

With a mass of ~1.6 x 107 Daltons and com- posed of approximately 2700 proteins, bacteriophage M13 has been employed as a molecular scaffold in bionanomaterials fabrication. In order to extend the versatility of M13 in this area, residue-specific unnatural amino acid incorporation was employed to successfully display azide functionalities on specific solvent-exposed positions of the pVIII major sheath protein of this bacteriophage. Employing a combination of engineered mutants of the gene coding for the pVIII protein, the methionine (Met) analog, L-azidohomoalanine (Aha), and a suitable Escherichia coli Met auxotroph for phage pro- duction, conditions were developed to produce M13 bacteri- ophage labeled with over 350 active azides (estimated by fluorescent dye labeling utilizing a strain-promoted azide- alkyne cycloaddition) and capable of azide-selective attach- ment to 5 nm gold nanoparticles as visualized by transmis- sion electron microscopy. The capability of this system to undergo dual labeling utilizing both chemical acylation and bioorthogonal cycloaddition reactions was also verified. The above stratagem should prove particularly advantageous in the preparation of assemblies of larger and more complex molecular architectures based on the M13 building block.