Demonstration by mass spectrometry that purified native Treponema pallidumrare outer membrane protein 1 (Tromp1) has a cleaved signal peptide

Purified native Tromp1 was subjected to mass spectrometric analysis in orde r to determine conclusively whether this protein possesses a cleaved or unc leaved signal peptide. The molecular masses of Tromp1, three Treponema pall idum lipoproteins, and a bovine serum albumin (BSA) control were determined by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry, The molecular masses of all of the T, pallidum lipoprote ins and BSA were,within 0.7% of their respective calculated masses, The mol ecular mass of Tromp1 was 31,510 Da, which is consistent with a signal-less form of Tromp1, given a calculated mass of unprocessed Tromp1 of 33,571 Da , a difference of 2,061 Da (a 6.5% difference). Purified native Tromp1 was also subjected to MALDI-TOF analysis in comparison to recombinant Tromp1 fo llowing cyanogen bromide cleavage, which further confirmed the identity of Tromp1 and showed that native Tromp1 was not degraded at the carboxy termin us. These studies confirm that Tromp1 is processed and does not contain an uncleaved signal peptide as previously reported.