FILM ARCHITECTURE IN BIOMOLECULAR ASSEMBLIES - EFFECT OF LINKER ON THE ORIENTATION OF GENETICALLY-ENGINEERED SURFACE-BOUND PROTEINS

This contribution presents strategies for the optimization of supramol ecular architecture aimed at controlling the organization of biomolecu les at solid surfaces. Myoglobin, modified by site-directed mutagenesi s to include a unique cysteine residue, is selectively chemisorbed to self-assembled haloalkylsilylated silica surfaces of varying n-alkyl c hain length (n = 2, 3, 8, 11, 15) to yield a series of surface-immobil ized recombinant protein assemblies. These supramolecular assemblies a re probed using tapping mode atomic force microscopy, wettability meas urements, Fourier transform infrared spectroscopy, and linear dichrois m spectroscopy to determine how the individual components comprising t hese structures (substrate, silane coupling layer, and protein) influe nce macromolecular protein ordering and stability. Surface roughness i s found to be a minor contributor in the determination of macromolecul ar ordering in these assemblies. In contrast, the nature of the underl ying silane self-assembled coupling layer is shown to strongly influen ce both the spatial and functional properties of the chemisorbed prote in. Silane coupling layers with short aliphatic chain lengths (n = 2, 3) produce highly trans-conformationally ordered structures upon which differential heme prosthetic group orientation cain be achieved. Long alkyl chain (n greater than or equal to 11) silane-derivatized surfac es also form ordered structures. The stability of myoglobin appended t o long chain aliphatic silylated surfaces is poor, however. The appare nt protein instability arises due to the increased hydrophobic charact er of these films. At intermediate alkyl chain length (n = 8), a confo rmationally disordered coupling layer with a high concentration of gau che defects is produced, regardless of the method of silane deposition or postdeposition processing. Chemisorption of myoglobin to the highl y disorganized assembly yields a random orientation of the protein.