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.
Stevens
Molly M
MM
Laboratory of Biophysics and Surface Analysis, School of Pharmacy, The University of Nottingham, Nottingham, NG7 2RD, United Kingdom.
Allen
Stephanie
S
Sakata
Jill K
JK
Davies
Martyn C
MC
Roberts
Clive J
CJ
Tendler
Saul J B
SJ
Tirrell
David A
DA
Williams
Philip M
PM
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