| Two-dimensional (2D) ordering of macromolecules is of particular interest due to their scientific importance as fundamental models for studying phase transitions and self-assembly. It is also a basis for the design and development of many biosensor and bioassay devices and engineered biomaterials. The ability to create and manipulate self-assembled structures of macromolecules requires a thorough understanding and control of the various intermolecular forces involved. In this study, we used the 2D streptavidin crystallization on lipid layers as a model system to study the effects of external variables such as lipid substrate, solution ionic strength, and polymeric precipitants on the crystallization process.; Streptavidin, a globular protein that shows exceptionally high specific binding affinity to biotin, has previously been crystallized on 2D lipid layers under relatively high ionic strength conditions. The 2D streptavidin crystals showed various characteristic morphologies and crystal symmetries with solution pH. At pH values below 5--6, corresponding to the isoelectric (pI) point, streptavidin forms characteristic needle-shaped crystals with high-density P1 space group. At pH values above the pI, the crystals take on the lowest-density square C222 lattice, while at pH values between 5 and 6, an intermediate crystal with P2 symmetry is found.; In this thesis, we first studied the crystallization of streptavidin on giant lipid bilayer vesicles. Once bound to the vesicle surface, streptavidin crystallized to form a rigid polycrystalline shell surrounding the vesicle. The change in solution pH changed the crystal morphology and growth pattern, and the vesicles were distorted into either roughened spheres or football-shaped ellipsoids. Micropipette aspiration of streptavidin-coated vesicles revealed the unique mechanical properties of the protein-lipid membrane. Permanent deformation was observed at low strain, while viscoelastic behavior was observed at higher deformation. Despite their extremely rigid appearance, the presence of an external polycrystalline shell does not increase the toughness of the vesicles beyond that of bare vesicles. The distinctive properties of the protein-lipid composite membrane can be traced back to the unique receptor-ligand interactions of the streptavidin-biotin system.; Since the giant vesicles were not stable in high ionic strength solutions, we used sucrose as our medium. While streptavidin is known not to crystallize in deionized water, it crystallized abundantly in sucrose with no added salt. This curious behavior allowed the study of the crystallization process in extremely low ionic strength solutions. It was found that the streptavidin crystals grown in giant vesicles only showed the lowest density C222 packing regardless of solution pH due to high electrostatic interactions. The gradual screening of electrostatic charges through the addition of NaCl allowed the transition between crystal forms to be carefully monitored, and a new crystal form was discovered in the process. In addition to sucrose, we also found that the addition of polymers to the 2D streptavidin system can induce, enhance, or inhibit the formation of streptavidin crystals. Under appropriate conditions, the addition of polymer can reduce the amount of protein required to crystallize by approximately 50%, and larger, higher quality crystals can be obtained. As a result, the inclusion of polymeric precipitants may serve as additional enhancement to the traditional 2D protein crystallization. |
