Much current interest in nanoscience focuses on the incorporation of biomolecules as functional components of nano-scale devices. Motors and sensors made of nucleic acids or proteins can perform work or transduce signals, through the exploitation and enhancement of their natural binding modes and functions. Full utilization of these native properties requires selective methods for the modification of biomolecules with organic or inorganic device components. In this work, I explore the use of actin as a nanoscale "track" and periplasmic binding proteins as reagentless molecular sensors.

Chapter 2 focuses on methods for the incorporation of actin fibers into potential device settings through the use of actin polymerization initiators that could be attached to surfaces. The small molecule jasplakinolide and a mutant formin protein have been investigated for this use. A diazonium reaction with 4-aminoacetophenone was used to introduce a ketone-functionality onto the β-tyrosine substituent of jasplakinolide. The identity of the product of this reaction was confirmed by MS/MS analysis, and its ability to nucleate actin fiber growth was explored. It was observed that the ketone-bearing jasplakinolide lost all actin nucleation activity. Next, a site-specific bioconjungation reaction was used to transaminate the N-terminus of a protein construct containing the formin homology 2 domain. This reaction introduced an aldehyde that was coupled with a hydroxylamine functionalized biotin.

In Chapter 3, periplasmic binding proteins (PBPs) were evaluated for use as fluorescence resonance energy transfer (FRET) based molecular sensors. To this end, PBPs from T. maritima and E. coli were labeled site-selectively with a pair of dyes suitable for FRET. One dye was installed via an N-terminal transamination, and an engineered cysteine was used to attach the second dye. Transfer of energy from the donor to the acceptor was observed, although the signal changes upon ligand binding were modest. The versatility of this double-modification strategy was also exploited for the surface attachment of dye-labeled glutamine binding protein.