Ultimate 3000 rslc system

All ESI–MS-measurements were performed on a Dionex Ultimate 3000 RSLC system using an Aeris Widepore XB-C8, 150 × 2.1 mm, 3.6 µm dp column (Phenomenex, USA). Separation of 1 µL sample was achieved by a linear gradient from (A) H₂O + 0.1% formic acid (FA) to (B) ACN + 0.1% FA at a flow rate of 300 µL/min and 45 °C. The gradient was initiated by a 0.5 min isocratic step at 2% B, followed by an increase to 75% B in 10 min to end with a 3 min step at 75% B before re-equilibration with initial conditions. UV spectra were recorded by a DAD in the range from 200 to 600 nm. The LC flow was split to 75 µL/min before entering the maXis 4G hr-ToF mass spectrometer (Bruker Daltonics, Bremen, Germany), using the standard Bruker ESI source. In the source region, the temperature was set to 200 °C, the capillary voltage was 4000 V, the dry-gas flow was 5.0 L/min and the nebulizer was set to 1.0 bar. Mass spectra were acquired in positive ionization mode ranging from 600 to 1800 m/z at 2.5 Hz scan rate. Protein masses were deconvoluted by using the Maximum Entropy algorithm (Copyright 1991–2004 Spectrum Square Associates, Inc.).

All ESI-UPLC-HRMS measurements were performed on a Dionex (Germering, Germany) Ultimate 3000 RSLC system coupled with a maXis4G Q-ToF mass spectrometer using an Aeris Widepore XB-C8 column (3.6 µm, 150 × 2.1 mm, Phenomenex) column. Separation of 1 µL sample was achieved by a multistep gradient from (A) H₂O + 0.1 % FA to (B) ACN + 0.1 % FA at a flow rate of 300 µL/min and 45 °C. Chromatographic conditions were as follows: 0–1 min, 2 % B; 1-11 min, 2–75 % B; 11–14 min, 75% B; 14-14.35 min, 75-2% B; 14.35-15.35 min, 2 % B. UV spectra were recorded by a DAD in the range from 200 to 600 nm. The LC flow was split to 75 µL/min before entering the maXis 4G hr-ToF mass spectrometer (Bruker Daltonik, Bremen, Germany) using the standard Bruker Apollo II ESI source. In the source region, the temperature was set to 200 °C, the capillary voltage was 4000 V, the dry-gas flow was 5.0 L/min and the nebulizer was set to 1.0 bar. Mass spectra were acquired in positive ionization mode ranging from 150 to 2500 m/z at 2 Hz scan rate. Protein masses were deconvoluted by using the Maximum Entropy algorithm in DataAnalysis 5.3 (Bruker Daltonik GmbH).