Demonstration by mass spectrometry that purified native Treponema pallidum rare outer membrane protein 1 (Tromp1) has a cleaved signal peptide.
Purified native Tromp1 was subjected to mass spectrometric analysis in order to determine conclusively whether this protein possesses a cleaved or uncleaved signal peptide. The molecular masses of Tromp1, three Treponema pallidum 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 lipoproteins and BSA were within 0.7% of their respective calculated masses. The molecular 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 following cyanogen bromide cleavage, which further confirmed the identity of Tromp1 and showed that native Tromp1 was not degraded at the carboxy terminus. These studies confirm that Tromp1 is processed and does not contain an uncleaved signal peptide as previously reported.
Blanco
D R
DR
Department of Microbiology, Immunology, and Molecular Genetics, University of California at Los Angeles, Los Angeles, California 90095, USA. DBLANCO@microimmun.medsch.ucla.edu
Whitelegge
J P
JP
Miller
J N
JN
Lovett
M A
MA
D001425:Bacterial Outer Membrane Proteins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins; D001483:Base Sequence; D017931:DNA Primers; D018272:Porins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D021382:Protein Sorting Signals / Q000032:analysis* / Q000378:metabolism; D011994:Recombinant Proteins / Q000032:analysis; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014210:Treponema pallidum / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D017931:DNA Primers; D018272:Porins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; C093836:Tromp1 protein, Treponema pallidum