pH-dependent behavior of surface-immobilized artificial leucine zipper proteins.
The coiled-coil protein motif occurs in over 200 proteins and has generated interest for a range of applications requiring surface immobilization of the constituent peptides. This paper describes an investigation of the environment-responsive behavior of a monolayer of surface-immobilized artificial proteins, which are known to assemble to form coiled-coil structures in bulk solution. An extended version of the quartz crystal microbalance (QCM-D) and surface plasmon resonance (SPR) are independently employed to characterize the adsorption of the proteins to a gold surface. The data suggest that the molecules arrange in a closely packed layer orientated perpendicular to the surface. QCM-D measurements are also employed to measure pH-induced changes in the resonant frequency (f) and the energy dissipation factor (D) of a gold-coated quartz crystal functionalized with the formed monolayer. Exposure of the protein monolayer to a pH 4.5 solution results in a shift of 43 Hz in f and a shift of -0.7 x 10(-6) in D as compared to pH 7.4. In contrast, increasing the pH to 11.2, results in f and D shifts of -17 Hz and 0.6 x 10(-6), respectively. The magnitude of the observed shifts suggests that the proteins form a rigid layer at low pH that can be hydrated to a fluid layer as the pH is increased. These observations correlate with spectroscopic changes that indicate a reduction in the helical content of the protein in bulk solutions of high pH.
Laboratory of Biophysics and Surface Analysis, School of Pharmacy, The University of Nottingham, Nottingham, NG7 2RD, United Kingdom.
Molly M
MM
Stevens
Stephanie
S
Allen
Jill K
JK
Sakata
Martyn C
MC
Davies
Clive J
CJ
Roberts
Saul J B
SJ
Tendler
David A
DA
Tirrell
Philip M
PM
Williams
D000327:Adsorption; D015374:Biosensing Techniques / Q000379:methods*; D020099:Coated Materials, Biocompatible / Q000737:chemistry*; D006046:Gold / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D016350:Leucine Zippers; D008565:Membrane Proteins / Q000737:chemistry*; D036103:Nanotechnology; D011485:Protein Binding; D011791:Quartz; D020349:Surface Plasmon Resonance; D013499:Surface Properties; D013997:Time Factors
D020099:Coated Materials, Biocompatible; D008565:Membrane Proteins; D011791:Quartz; D006046:Gold