Ultraflex 1 tof tof mass spectrometer

In all, 1 ml of each sterile filtrated bacterial supernatant was acidified with 0.4 ml of 1% trifluoric acid (TFA; Applied Biosystems, Warrington, UK). The solution was loaded onto an activated C18-SPE-cartridge (100 mg; Macherey-Nagel, Düren, Germany); the retained peptides were washed with 0.1% TFA and eluted with 80% acetonitrile (Roth GmbH, Karlsruhe, Germany) in 0.1% TFA. The eluate was lyophilised overnight, redissolved in 20 µl 0.1% TFA, and 1 µl of this sample was mixed with 1 µl matrix solution (2, 5-dihydroxybenzoic acid, DHB; 5 mg/ml; Sigma, Darmstadt, Germany), and methylendiphosphonic acid (MDPA; 5 mg/ml; Fluka, Steinheim, Germany) in 0.1% TFA) for MS analysis. MALDI-TOF-MS was performed on an Ultraflex I TOF/TOF mass spectrometer (Bruker Daltonics, Bremen, Germany) equipped with a nitrogen laser and a LIFT-MS/MS facility. The instrument was operated in the positive-ion reflectron mode acquiring 200–400 single spectra per sample for sum spectra. For data processing and instrument control the Compass 1.3 software package consisting of FlexControl 2.4, FlexAnalysis 3.0, BioTools 3.0 was used.

MALDI-TOF MS was performed using an UltraflexI TOF/TOF mass spectrometer (Bruker Daltonics, Bremen, Germany) equipped with a nitrogen laser and a LIFT-MS/MS facility. The instrument was operated in the positive-ion reflectron mode using 2,5-dihydroxybenzoic acid (Sigma) in 50% ACN/1% phosphoric acid as matrix. Acquired sum spectra consist of 200–400 single spectra. For data processing and instrument control the Compass 1.1 software package consisting of FlexControl 2.4, FlexAnalysis 2.4, and ProteinScape 3.0 was used.

Matrix-assisted laser-desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) was performed on an Ultraflex I TOF/TOF mass spectrometer (Bruker Daltonics, Bremen, Germany) equipped with a nitrogen laser. The instrument was operated in the positive-ion reflectron mode using 2,5-dihydroxybenzoic acid (5 mg/mL; Sigma), and methylenediphosphonic acid (5 mg/mL; Fluka) in 0.1% TFA as matrix. Sum spectra consisting of 200–400 single spectra were acquired. For data processing and instrument control, the Compass 1.4 software package consisting of FlexControl 3.4, FlexAnalysis 3.4, BioTools 3.2 was used. The correlation of the sequence of USCTX with the observed m/z-values was analyzed with BioTools, allowing one missed cleavage. The oxidation and crosslinking of cysteines were set as variable modifications. The obtained MALDI data are given in Supplementary Figure S1.