Amphiphilic diblock copolymers for molecular recognition

In this thesis, the synthesis and the characterization of poly(butadiene)-blockpoly(
ethylene oxide) copolymers with terminal Me2+-NTA groups (copper or nickel) is
described for the first time. A convenient “one-pot” procedure that allows control over the
individual block lengths of the copolymer and the end-group functionalization was
successfully established.
The formation of the metal-polymer complex has been confirmed by EPR and UV/VIS
spectroscopy. Mixing of the Ni2+-NTA polymers with the corresponding non functionalized
block copolymers at a concentration of 10 mol% does not affect the self-assembly behavior
of the mixtures, i.e., in dilute aqueous solutions the polymer mixtures aggregate to vesicular
structures (metal-doped vesicles) with identical size distribution as the non functionalized
block copolymer vesicles. Vesicles were characterized by dynamic light scattering, static
light scattering, small angle X-ray scattering and zeta potential. All measurements led to the
conclusion that hollow spheres, i.e. vesicles, with a narrow size distribution and a negative
surface potential were generated. Moreover different vesicle shapes as “necklace pearls”,
“wormlike micelles” and “spermasomes” can be attributed to different salt solutions or
buffers of defined concentrations which suggests a control of morphology.
The accessibility of the metal sites at the surface of such vesicles has been tested using
fluorescence correlation spectroscopy. The model proteins His10-MBP-FITC and His6-EGFP
bind selectively to the Me2+-NTA groups exposed at the surface of the vesicles. While the
choice of the buffer significantly influenced the fractions of protein-vesicle conjugates, the
interactions of Cu2+- and Ni2+-NTA groups with both His-tagged proteins showed similar
values. It should be noted that the experimentally determined dissociation constants of the
Me2+-His-Tag complexes were found to be in good agreement with literature data on Ni-
NTA functionalized liposomes14, indicating that the polymer brushes at the polymer vesicle
surface only slightly interfere with the binding of the proteins.
Fluorescence Microscopy was used to visualize the binding of the fluorescent proteins to
the functionalized vesicles and images of vesicles with a fluorescent corona were taken.
Additionally, atomic force microscopy clearly demonstrated that the polymer adsorbs in
an oriented manner on highly oriented pyrolytic graphite surfaces and is able to induce a 2D
protein crystallization when Ni-NTA functionalized polymer was used.
We believe that these metal-functionalized polymeric membranes have a large potential
for the selective immobilization and alignment of proteins at vesicle/planar membrane
surfaces. In particular, the high flexibility and compressibility of block copolymer membranes
and monolayers could open new possibilities for inducing a 2D protein crystallization. The
high cohesion and robustness of block copolymer membranes make them rather insensitive
toward mechanical shear or the presence of detergents, increasing their potential utility. In
this context, it should also be noted that the pendant double bonds of the poly(butadiene)
blocks can be covalently cross-linked, thus freezing the self-assembled structures and
providing additional stabilization.