Size-dependent control of the binding of biotinylated proteins to streptavidin using a polymer shield.
Many medical and biotechnological processes rely on controlling and manipulating the molecular-recognition capabilities of proteins. This can be achieved using small molecules capable of competing for protein binding or by changing environmental parameters that affect protein structure and hence binding. An alternative is provided by stimuli-responsive polymers that change reversibly from a water-soluble expanded coil to a water-insoluble collapsed globule upon small changes in temperature, pH or light intensity: when attached to proteins in the vicinity of their binding sites, they reversibly block and release small ligands. Here we show how this approach can be extended to achieve size-selective binding of large, macromolecular ligands. We use the thermally responsive polymer poly(N,N-diethylacrylamide) (PDEAAm), and attach it to the protein streptavidin approximately 20 A from the binding site for biotinylated proteins. Below the lower critical solution temperature of PDEAAm, the polymer is in its extended state and acts as a 'shield' to block the binding of large biotinylated proteins; above this temperature, it collapses and exposes the binding site, thereby allowing binding. We find that the degree of shielding depends on both the size of the biotinylated protein and the size of PDEAAm, suggesting that 'smart' polymer shields could be tailored to achieve a wide range of size-dependent ligand discrimination for use in affinity separations, biosensors and diagnostics technologies.
Ding
Z
Z
Department of Bioengineering, University of Washington, Seattle 98195, USA.
Fong
R B
RB
Long
C J
CJ
Stayton
P S
PS
Hoffman
A S
AS
D000178:Acrylamides; D001710:Biotin / Q000737:chemistry / Q000378:metabolism*; D010316:Particle Size; D011108:Polymers / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D019809:Streptavidin / Q000737:chemistry / Q000378:metabolism*
D000178:Acrylamides; D011108:Polymers; C428028:poly(N,N-diethylacrylamide); D001710:Biotin; D019809:Streptavidin