RAPID SDS-GEL CAPILLARY ELECTROPHORESIS FOR THE ANALYSIS OF RECOMBINANT NADP(-DEPENDENT FORMATE DEHYDROGENASE DURING EXPRESSION IN ESCHERICHIA-COLI-CELLS AND ITS PURIFICATION())

The level of expression in Escherichia coli cells and different steps of purification of the recombinant NADP(+)-dependent formate dehydroge nase (EC 1.2.1.2, FDH) from bacterium Pseudomonas sp.101 was analyzed by rapid SDS-Gel capillary electrophoresis (SDS-Gel CE) and compared w ith SDS polyacrylamide gel electrophoresis (SDS PAGE). First standard proteins were separated in the short capillary and the calibration cur ve generated, then fractions taken during the fermentation and purific ation process were analyzed. The main advantages of SDS-Gel CE are sho rt analysis time, high sensitivity, the possibility to quantify protei ns at different ultraviolet wavelength, and small injection volumes. T he data for each step of the fermentation process and during the purif ication were controlled by spectrophotometric analysis of enzyme activ ity and protein concentration as well as standard SDS PAGE. The molecu lar mass of the purified FDH was determined as 44 078 Da by matrix-ass isted laser desorption/ionisation time of flight mass spectrometry (C) 1997 Elsevier Science B.V.