BETA-LACTAMASE RAGGED ENDS DETECTED BY ELECTROSPRAY MASS-SPECTROMETRYCORRELATES POORLY WITH MULTIPLE BANDING ON ISOELECTRIC-FOCUSING

Purified preparations of TEM-2, P99, Bacillus cereus I and B. cereus I I beta-lactamases were examined by electrospray (ES) mass spectrometry . The ES mass spectra of the B. cereus enzymes revealed the presence o f four to five components of different mass, corresponding to the loss of different numbers of N-terminal amino acids (ragged ends). The ES mass spectra of both TEM-2 and P99 consisted of a single component wit h no evidence of ragged ends. All four beta-lactamase preparations wer e visualized on isoelectric focusing (IEF) gels stained with nitrocefi n to investigate a possible correlation between IEF patterns and ragge d ends. Multiple banding patterns were seen with each beta-lactamase p reparation. Although these may correlate with the presence of ragged e nds in the two B. cereus preparations, the satellite bands seen with P 99 and TEM-2 were not associated with differences detected by ES mass spectrometry. In this study we have shown for the first time that beta -lactamase satellite bands seen on IEF are not always associated with ragged ends. Furthermore, we have illustrated the use of ES mass spect rometry to characterize the extent of ragged end formation in protein samples. This is of particular significance if the sample is required for detailed biochemical or crystallography experiments.