ELECTROSPRAY-IONIZATION MASS-SPECTROMETRY OF GENETICALLY AND CHEMICALLY-MODIFIED BACTERIORHODOPSINS

A simple method for the preparation of bacteriorhodopsin for mass spec trometry is described, consisting of precipitation of the sample, remo val of lipids by washing the sample, and dissolving it in an acidic mi xture of organic solvents. Examples demonstrate the method's suitabili ty for the analysis of genetically and chemically modified bacteriorho dopsins. The observed molecular masses are within 0.01% in accordance with the theoretical masses of the wild-type protein and modification products. This allows the detection of modifications accompanied by a mass change greater than 2 Da. Thus, only 9 of 380 possible binary ami no acid exchanges are not amenable to analysis. Also chemical modifica tions of bacteriorhodopsin can be followed since it is possible to cha racterize reaction products and to approximate yields. However, second ary reactions like hydrolysis in the reaction medium or in the acid so lvent used for sample preparation must be considered. (C) 1996 Academi c Press, Inc.