A STUDY OF THE THERMAL-DENATURATION OF RIBONUCLEASE-S BY ELECTROSPRAY-IONIZATION MASS-SPECTROMETRY

The thermal stability of ribonuclease S (RNase S), an enzymatically ac tive noncovalent complex composed of a 2166-u peptide (S-peptide) and a 11,534-u protein (S-protein), was investigated by electrospray ioniz ation mass spectrometry (ESI-MS) and capillary electrophoresis ESI-MS (CE-ESI-MS). The intensities of peaks corresponding to the RNase S com plex were inversely related to both the applied nozzle-skimmer (or cap illary-skimmer) voltage bias in the atmosphere-vacuum interface and th e temperature of the RNase S solution. By using a heated metal capilla ry-skimmer interface and a room temperature solution of RNase S, the i ntensities of RNase S molecular ion peaks were observed to decrease wi th increasing metal capillary temperature. Mass spectrometric studies with both the nozzle-skimmer and capillary-skimmer interface designs a llowed determination of phenomenological enthalpies for dissociation o f the RNase S complex in both solution and for the electrosprayed micr odroplet-gas phase species. Intact RNase S complex could also be detec ted with CE-ESI-MS separations by using a 10-mM ammonium bicarbonate ( pH 7.9) solution as the electrophoretic buffer. These studies provide new insights into the stability of multiply charged noncovalent comple xes in the gas phase and the mass spectrometric conditions required fo r such studies, and suggest that information regarding solution proper ties can be obtained by ESI-MS.