Trypsinization and Tandem Mass Spectrometry

Trypsinization of proteins was performed as previously described^{52 (link),53 (link)}. Tandem MS (MS/MS) analysis was performed using an LTQ Orbitrap ELITE ETD mass spectrometer (Thermo Fisher Scientific). The mass spectrometer was operated in data-dependent acquisition mode, in which MS acquisition with a mass range of 400 to 1000 m/z was automatically switched to MS/MS acquisition under the control of the Xcalibur software (Thermo Fisher Scientific). The top four precursor ions in the MS scan were selected using Orbitrap, with a resolution of R = 240,000, and the ions in subsequent MS/MS scans were analyzed with an ion trap in automated gain control (AGC) mode, in which AGC values were 1 × 10⁶ and 1.00 × 10⁴ for full MS and MS/MS, respectively. Electron transfer dissociation (ETD) was used for fragmentation.

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Okuda H., Miyamoto R., Takahashi S., Kawamura T., Ichikawa J., Harada I., Tamura T, & Yokoyama A. (2022). RNA-binding proteins of KHDRBS and IGF2BP families control the oncogenic activity of MLL-AF4. Nature Communications, 13, 6688.

Publication 2022

LTQ Orbitrap ELITE ETD mass spectrometer Xcalibur software

Electron transfer Ms ms Proteins Scans

Corresponding Organization : National Institute of Technology, Tsuruoka College

Other organizations : The University of Tokyo, Yokohama City University

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Variable analysis

independent variables

Trypsinization of proteins

dependent variables

Tandem MS (MS/MS) analysis

control variables

Mass spectrometer operation in data-dependent acquisition mode
Mass range of 400 to 1000 m/z for MS acquisition
Orbitrap resolution of R = 240,000 for MS scans
Automated gain control (AGC) values of 1 × 10^6 and 1.00 × 10^4 for full MS and MS/MS, respectively
Electron transfer dissociation (ETD) for fragmentation

controls

No positive or negative controls were explicitly mentioned in the given information.

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