Observation of charge state and conformational change in immobilized protein using surface plasmon resonance sensor

Behaviors of proteins immobilized on a solid surface were investigated usin g BIACORE, a biosensor utilizing surface plasmon resonance. This sensor is usually used for analyzing binding events during biomolecular interactions. Here we propose a novel use of this sensor to monitor two kinds of intramo lecular changes in immobilized proteins. Several proteins were covalently a ttached to dextran chains on the sensor surface in the flow cell and were t hen exposed to a series of buffers with varying pH. Signal changes derived from changes of refractive index around the sensor surface were detected du ring and after the exposure to each of these buffers, which we denoted as i n situ values and postvalues, respectively. The in situ value reflects the behavior of immobilized proteins in these buffers and was revealed to have a correlation with total charge state of the proteins, while the postvalue reflects how immobilized proteins react after the exposure and was suggeste d to represent the degree of conformational changes of the proteins, This m ethod is expected to be applicable to various analyses and can provide us w ith new information about the behavior of proteins on solid phase. (C) 2001 Academic Press.