AVS 62nd International Symposium & Exhibition
    Biomaterial Interfaces Monday Sessions
       Session BI+AS-MoA

Paper BI+AS-MoA2
Controlled Molecular Mechanisms of Engineered Solid Binding Proteins on Surfaces

Monday, October 19, 2015, 2:40 pm, Room 211D

Session: Characterization of Biological and Biomaterials Surfaces (2)
Presenter: Christopher So, National Research Council postdoc cited at Naval Research Laboratory
Authors: C. So, National Research Council postdoc cited at Naval Research Laboratory
S. Walper, US Naval Research Laboratory
R. Stine, Nova Research
D.E. Barlow, US Naval Research Laboratory
K. Wahl, US Naval Research Laboratory
Correspondent: Click to Email
Persistent and uncontrolled aggregation of proteins at surfaces remains a major challenge for biocompatibility, fouling, and biosensing. To fully realize the rich properties of proteins at interfaces, a critical link between displayed protein sequence and surface assembly mechanisms is required. Here we use rational protein mutations combined with in situ microscopy and spectroscopy methods to demonstrate that manipulation of solid binding and intermolecular interactions by proteins can dictate their surface behavior and induce nanostructure formation. We use streptavidin (SA) as a robust scaffold to control the density and localization of aromatic residues, expected to interact with surfaces such as graphite and graphene through pi-bonding. The surface adapted SAs are generated by placing aromatic side chains of varying polarity (Phenylalanine, Tyrosine, Tryptophan) along three putative permissive sites in a coplanar arrangement. The effects of these mutations on bulk solution structure, surface-associated structure, as well as surface affinity, orientation and spatial organization are studied in situ using attenuated total reflectance (ATR) infrared spectroscopy (IR) with linear polarization (LP), fluid-mode atomic force microscopy (AFM), and circular dichroism (CD). We have found that our simple modifications to mSA have little effect on the solution state of the protein, while having a pronounced effect on affinity and secondary structure in the adsorbed state. Through fabricating graphene-coated ATR-IR prisms, we find that unmodified mSA exhibits an ordered beta sheet structure at surfaces, while tryptophan modifications to mSA (Trp-mSA) induces a more disordered structure. We quantify by temporal ATR-IR spectra a ca. 4.5x enhancement in sticking probability for Trp-mSA over mSA to graphene. Fluid-mode AFM studies on graphite support a surface-mediated coarsening mechanism: while mSA forms no obvious surface structures, Trp-mSA aggregates and forms islands 10-50 nm in size over the course of an hour. Such disordered SA aggregates provide high affinity sites for slow lateral island growth processes, giving rise to a bi-modal exponential adsorption curve for Trp-mSA but absent in mSA. Ultimately, defining the molecular basis of protein self-assembly and the impact of displayed chemistries at liquid-solid interfaces will enable rationally designed biological surface coatings and engineered biointerfaces with tailorable functionalities.