Binding kinetics between soluble β-Gal and anti-β-Gal immobilized on chemically nanopatterned surfaces with fractal topography

Abstract

The present work was aimed at providing experimental support to the correlation between topographic and kinetic dimensions of protein function we described previously. Through electronic and colloidal nanolitographic techniques we produced surfaces exhibiting a fractal-like pattern with a pre-determined topographic dimension of domains capable to bind proteins in a covalent manner (AbAntiβ -Gal). These surfaces were used as sensors to reversibly bind β-Gal and enabled the kinetic study of AbAnti β-Gal -β-Gal complex formation by surface plasmon resonance (SPR) spectroscopy. Compared with the behavior of control sensors (homogeneous topography), the Ag-Ab binding kinetics in sensors produced by nanolithography showed higher capacity and broader dispersion of binding sites characterized by a more diffuse attractor in the kd vs. Kd phase space. Our results contributed to the development of a nano-biosensor for β-Gal, to a strategy to enhance the sensitivity of a SPR sensor and to endorse the cross-talk between supramolecular structure and dynamics in heterogenous systems. Additionally, a catalytic activity could be measured for the first time by a SPR based method where the insoluble product of a β-Gal catalyzed reaction interfered with the evanescent field on the sensor´s surface.