Sterol carrier protein 2 (SCP2) has been investigated by nearly native electrospray ionisation mass spectrometry in the presence of long chain fatty acyl CoAs (LCFA-CoAs) and carnitine derivatives of equivalent fatty acid chain length (LCFA-carnitines). Four SCP2 constructs were compared to examine the influence of the N-terminal presequence and the C-terminal peroxisomal targeting signal on ligand binding. Removal of N- or C-terminal residues did not influence ligand binding. The observation that LCFA-CoAs are high affinity ligands for SCP2 was confirmed, while LCFA-carnitines were demonstrated for the first time not to interact with SCP2. LCFA-CoAs formed non-covalent complexes with SCP2 of 2:1 and 1:1 stoichiometry, which could be dissociated by elevating the energy of the ions upon entrance to the mass spectrometer. A fluorescence-competition assay using Nile Red butyric acid confirmed the mass spectrometric observations in solution. The physiological significance of the lack of LCFA-carnitine binding by SCP2 is discussed. (c) 2007 Elsevier Inc. All rights reserved.
Stanley, W., Versluis, K., Schultz, C., Heck, A. J. R., & Wilmanns, M. (2007). Investigation of the ligand spectrum of human sterol carrier protein 2 using a direct mass spectrometry assay. Archives of Biochemistry and Biophysics, 461(1), 50-58. https://doi.org/10.1016/j.abb.2007.02.026