Lipid Binding Spectrum of OmpF Trimer

Lipid stocks of 1,2-dimyristoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (DMPG), 1,2-dipalmitoyl-snglycero-3-phospho-(1′-rac-glycerol) (DPPG) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (POPG) were prepared from powder (Avanti Polar Lipids Inc., Alabama USA) at concentrations of 10 mg/mL using the previously published procedure ^{14 (link)}. Lipid stocks were diluted 50 fold in 1 % (w/v) β-OG, 200 mM ammonium acetate and added to OmpF in an appropriate ratio for multiple lipid species to be bound. For multiple lipid binding an approximately equimolar mixture of all three lipid species was prepared. The spectrum of concomitant binding of DMPG, DPPG and POPG to the OmpF trimer was acquired at an increased transient time of 128 ms (35,000 resolution at m/z 200).

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Gault J., Donlan J.A., Liko I., Hopper J.T., Gupta K., Housden N.G., Struwe W.B., Marty M.T., Mize T., Bechara C., Zhu Y., Wu B., Kleanthous C., Belov M., Damoc E., Makarov A, & Robinson C.V. (2016). High-resolution mass spectrometry of small molecules bound to membrane proteins. Nature methods, 13(4), 333-336.

Publication 2016

Ammonium acetate Dmpg Dppg Glycerol Lipid M 200 Powder Transient

Corresponding Organization :

Other organizations : University of Oxford, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Thermo Fisher Scientific (Germany)

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Variable analysis

independent variables

Lipid species (DMPG, DPPG, POPG)
Lipid concentration (10 mg/mL)
Lipid dilution (50-fold in 1% (w/v) β-OG, 200 mM ammonium acetate)
Mixing ratio of lipid species for multiple lipid binding

dependent variables

Concomitant binding of DMPG, DPPG, and POPG to the OmpF trimer
Spectrum acquired at an increased transient time of 128 ms (35,000 resolution at m/z 200)

control variables

Previously published procedure for lipid stock preparation (¹⁴)
Presence of OmpF in the binding experiment

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

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