MassIVE MSV000084963

Partial Public PXD017565

ARIH2 is a Vif-dependent regulator of CUL5-mediated APOBEC3G degradation in HIV infection: SRM-CUL5

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Description

SRM assays were generated for selected interactors of CUL5. SRM assay generation was performed using Skyline. For all targeted proteins, proteotypic peptides and optimal transitions for identification and quantification were selected based on the Skyline spectral library generated from the shotgun MS experiments (MSV000084855). For each protein, 2-5 peptides were selected based on intensity, peptide length as well as chromatographic performance. For each peptide the 4 best SRM transitions were selected based on intensity and peak shape. Digested peptide mixtures were analyzed by LC-SRM on a Thermo Scientific TSQ Quantiva MS system equipped with a Proxeon. Easy nLC 1200 ultra high-pressure liquid chromatography and autosampler system. Samples were injected onto a C18 column (25 cm x 75 mm I.D. packed with ReproSil Pur C18 AQ 1.9 mm particles) in 0.1% formic acid and then separated with an 80 min gradient from 5% to 40% Buffer B (90% ACN/10% water/0.1% formic acid) at a flow rate of 300 nL/min. SRM acquisition was per- formed operating Q1 and Q3 at 0.7 unit mass resolution. For each peptide the best 4 transitions were monitored in a scheduled fashion with a retention time window of 4 min and a cycle time fixed to 2 seconds. Argon was used as the collision gas at a nominal pressure of 1.5 mTorr. Collision energies were calculated by, CE = 0.0348 * (m/z) + 0.4551 and CE = 0.0271 * (m/z) + 1.5910 (CE, collision energy and m/z, mass to charge ratio) for doubly and triply charged precursor ions, respectively. RF lens voltages were calculated by, RF = 0.1088 * (m/z) + 21.029 and RF = 0.1157 * (m/z) + 0.1157 (RF, RF lens voltage and m/z, mass to charge ratio) for doubly and triply charged precursor ions, respectively. The resulting data was analyzed with Skyline for identification and quantification of peptides. MSstats was used for statistical analysis. [doi:10.25345/C5BX0X] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: Host-virus interactions ; ARIH2 ; HIV ; vif ; CUL5 ; AP-MS ; APOBEC3G ; MassIVE.quant reviewed - Platinum

Contact

Principal Investigators:
(in alphabetical order) 		Nevan J. Krogan, University of California, San Francisco, United States
Submitting User: mnchoi

Publications

Hüttenhain R, Xu J, Burton LA, Gordon DE, Hultquist JF, Johnson JR, Satkamp L, Hiatt J, Rhee DY, Baek K, Crosby DC, Frankel AD, Marson A, Harper JW, Alpi AF, Schulman BA, Gross JD, Krogan NJ.
ARIH2 Is a Vif-Dependent Regulator of CUL5-Mediated APOBEC3G Degradation in HIV Infection.
Cell Host Microbe. 2019 Jul 10;26(1):86-99.e7. Epub 2019 Jun 25.

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